JP6884608B2 - Compositions and external preparations for skin - Google Patents
Compositions and external preparations for skin Download PDFInfo
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- JP6884608B2 JP6884608B2 JP2017046598A JP2017046598A JP6884608B2 JP 6884608 B2 JP6884608 B2 JP 6884608B2 JP 2017046598 A JP2017046598 A JP 2017046598A JP 2017046598 A JP2017046598 A JP 2017046598A JP 6884608 B2 JP6884608 B2 JP 6884608B2
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Landscapes
- Coloring Foods And Improving Nutritive Qualities (AREA)
- Cosmetics (AREA)
- Medicines Containing Material From Animals Or Micro-Organisms (AREA)
- Medicines Containing Plant Substances (AREA)
Description
本発明は、複数の天然物由来成分を有し、すぐれた生理活性及び生体安全性を有する機能性素材及びこれを配合してなる皮膚(頭皮も含む)外用剤及び美容又は健康増進用の飲食品である。 The present invention is a functional material having a plurality of natural product-derived components and having excellent physiological activity and biosafety, a skin (including scalp) external preparation containing the same, and food and drink for beauty or health promotion. It is a product.
近年、細胞の老化現象や外的因子(例えば、紫外線、大気汚染物質や環境ホルモン等の化学物質、花粉等のアレルギー物質、環境ストレス等)による細胞へのダメージに関する研究が行われ、様々な細胞の老化現象や、細胞の損傷及び修復に関するメカニズムが解明されている。 In recent years, research has been conducted on cell aging and damage to cells caused by external factors (for example, ultraviolet rays, chemical substances such as air pollutants and environmental hormones, allergens such as pollen, environmental stress, etc.), and various cells have been studied. The mechanism of senescence and cell damage and repair has been elucidated.
例えば、皮膚細胞に関しては、加齢に伴う細胞増殖・分化の不活化、ホルモン分泌の低下、皮膚を構成する細胞外マトリックス成分(コラーゲン、ラミニン等)の量的低下等の内的要因と、太陽光(紫外線)に誘発される活性酸素、大気汚染物質や環境ホルモン等の化学物質、花粉等のアレルギー物質、環境ストレス等の外的要因とが複雑に絡み合って、老化現象(シワ、タルミ等)や肌荒れ、色調の変化が生じることが知られている。 For example, regarding skin cells, internal factors such as inactivation of cell proliferation / differentiation with aging, decrease in hormone secretion, quantitative decrease in extracellular matrix components (collagen, laminin, etc.) constituting the skin, and the sun Aging phenomenon (wrinkles, tarmi, etc.) due to complex intertwining of active oxygen induced by light (ultraviolet rays), chemical substances such as air pollutants and environmental hormones, allergens such as pollen, and external factors such as environmental stress. It is known that rough skin and changes in color tone occur.
さらに、外的要因である紫外線、化学物質、アレルギー物質は、生体内の細胞や組織にダメージを与えて生体成分を変質させたり、又は活性酸素を発生させたりする。これにより、細胞内に抗原を発生し、生体において炎症が生じる。さらには、上記外的要因が細胞内のメラニン色素の異常沈着を誘発して皮膚にシミ、ソバカス、肝斑等を生じさせる。 Furthermore, ultraviolet rays, chemical substances, and allergens, which are external factors, damage cells and tissues in the living body to alter biological components or generate active oxygen. As a result, an antigen is generated inside the cell, and inflammation occurs in the living body. Furthermore, the above-mentioned external factors induce abnormal deposition of intracellular melanin pigment, causing spots, freckles, liver spots and the like on the skin.
以上のような細胞の不健全化や老化を予防及び改善する目的で、従来、種々の活性成分が提案され、それら活性成分を配合した化粧品、飲食品及び医薬品が上市されている。例えば、ビタミンC、ビタミンE、カタラーゼ等の抗酸化剤;グリチルリチン酸又はその塩、アラントイン、トラネキサム酸等の抗炎症剤;各種紫外線吸収剤;α−ヒドロキシカルボン酸、胎盤抽出液、γ−アミノ−β−ヒドロキシ酪酸等の細胞賦活成分;コラーゲン、エラスチン、ヒアルロン酸又はその塩等の細胞外マトリックス成分;尿素等の保湿剤;アミノグアニジン等のタンパク質糖化抑制剤が挙げられる。また、皮膚のシミ、ソバカス、肝斑等の色素沈着の発生を抑制する物質としては、コウジ酸やリノール酸等が知られており、美白剤の有効成分として広く使用されている。 For the purpose of preventing and improving the unhealthyness and aging of cells as described above, various active ingredients have been conventionally proposed, and cosmetics, foods and drinks and pharmaceuticals containing these active ingredients have been put on the market. For example, antioxidants such as vitamin C, vitamin E and catalase; anti-inflammatory agents such as glycyrrhizic acid or salts thereof, allantin, tranexamic acid; various ultraviolet absorbers; α-hydroxycarboxylic acid, placenta extract, γ-amino- Cell-activating components such as β-hydroxybutyric acid; extracellular matrix components such as collagen, elastin, hyaluronic acid or salts thereof; moisturizers such as urea; protein glycation inhibitors such as aminoguanidine. Further, kojic acid, linoleic acid and the like are known as substances that suppress the occurrence of pigmentation such as skin stains, freckles and chloasma, and are widely used as active ingredients of whitening agents.
以上のように、従来、細胞の老化現象や不健全化のメカニズムに基づいて、細胞賦活剤、抗老化剤及び美白剤が提案されているが、生体に対する安全性、また、実際に生体への塗布又は服用に際しての有効性の観点で問題が存在する。従って、それら従来の問題点が改善された機能性素材が求められている。 As described above, cell activators, anti-aging agents and whitening agents have been conventionally proposed based on the mechanism of cell aging phenomenon and unhealthyness, but they are safe for living organisms and actually applied to living organisms. There is a problem in terms of effectiveness when applied or taken. Therefore, there is a demand for a functional material in which these conventional problems are improved.
本発明者らは、かかる従来技術の問題点に鑑みて、生体安全性の観点から天然物由来の新たな有効成分を見出すべく鋭意研究を行った。その結果、バラ科サクラ属に属するサクラの抽出物と、酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の抽出物又は酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の微生物により産生される微生物由来発酵代謝物を含む組成物が、すぐれた細胞賦活作用、抗酸化作用、コラーゲン合成促進作用、メラニン合成抑制及び脂肪蓄積抑制を有し、皮膚(頭皮も含む)外用剤や飲食品用の機能性素材として有用であることを見出した。 In view of the problems of the prior art, the present inventors have conducted diligent research to find a new active ingredient derived from a natural product from the viewpoint of biosafety. As a result, by the extract of Plum belonging to the genus Plum of the Rosaceae family and the extract of any one or more of yeast, aspergillus and lactic acid bacteria, or the microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. The composition containing the produced microbial-derived fermentation metabolite has excellent cell activation effect, antioxidant effect, collagen synthesis promoting effect, melanin synthesis suppression and fat accumulation suppression, and is used as an external preparation for skin (including scalp) and food and drink. We have found that it is useful as a functional material for goods.
従来、バラ科サクラ属に属するサクラの抽出物又は酵母抽出物が、皮膚生理活性を有することは、例えば、特許文献1〜6に開示されているが、サクラの抽出物と酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の抽出物又は酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の微生物により産生される微生物由来発酵代謝物とを含む組成物が、皮膚外用剤や飲食品用の機能性素材として有用であることについては、知られていなかった。
本発明は、バラ科サクラ属に属するサクラの抽出物と、酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の抽出物又は酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の微生物により産生される微生物由来発酵代謝物とを含有する組成物である。
また、本発明は、上記組成物を含む皮膚(頭皮も含む)外用剤又は飲食品であることを特徴とする。
The present invention relates to an extract of Plum belonging to the genus Plum of the Rosaceae family, an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite derived from a microorganism produced by.
The present invention is also characterized in that it is a skin (including scalp) external preparation or food or drink containing the above composition.
本発明は、バラ科サクラ属に属するサクラの抽出物と、酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の抽出物又は酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の微生物により産生される微生物由来発酵代謝物とを含有する組成物であって、当該組成物は、細胞賦活作用、抗酸化作用、コラーゲン産生促進、メラニン合成抑制及び脂肪蓄積抑制の相乗作用を有することから、細胞の健全化効果、紫外線等の外的要因による生体へのダメージ(酸化ダメージ及び炎症ダメージ)の予防及び改善効果、保湿効果、皮膚のターンオーバー改善効果、肌のシワ、タルミの予防及び改善効果、シミ、ソバカスの予防及び改善効果、皮脂抑制効果及び痩身効果を発揮する。これにより、本発明に係る組成物は皮膚外用剤や飲食品用の機能性素材として有用である。さらに、頭皮中のコラーゲンの産生を促進する効果も有することから、脱毛予防又は白髪予防用の機能性素材として有用である。 The present invention relates to an extract of cherry blossoms belonging to the genus Sakura of the family Rose family, an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism of any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite derived from a microorganism produced by the above, and the composition has a synergistic effect of cell activation, antioxidant action, promotion of collagen production, suppression of melanin synthesis and suppression of fat accumulation. , Cell health effect, prevention and improvement effect of damage to living body (oxidative damage and inflammatory damage) due to external factors such as ultraviolet rays, moisturizing effect, skin turnover improvement effect, prevention and improvement of skin wrinkles and melanin It exerts effects, prevention and improvement effects of spots and buckwheat, sebum suppression effect and slimming effect. As a result, the composition according to the present invention is useful as a functional material for external preparations for skin and foods and drinks. Furthermore, since it also has the effect of promoting the production of collagen in the scalp, it is useful as a functional material for preventing hair loss or gray hair.
以下、本発明の好ましい実施の形態について詳細に説明する。
本発明は、サクラの抽出物と、酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の抽出物又は酵母、麹菌及び乳酸菌のいずれか1種又は2種以上の微生物により産生される微生物由来発酵代謝物とを含有する組成物である。
Hereinafter, preferred embodiments of the present invention will be described in detail.
The present invention is derived from an extract of cherry blossoms and an extract of any one or more of yeast, aspergillus and lactic acid bacteria, or a microorganism produced by any one or more of yeast, aspergillus and lactic acid bacteria. It is a composition containing a fermentation metabolite.
本発明において、「サクラ」とは、バラ科サクラ属に属する植物であって、例えば、オオシマザクラ、オオヤマザクラ、カスミザクラ、ヤマザクラ(山桜)等のヤマザクラ(Prunus jamasakura)、ヤエザクラ、カンザン、フゲンゾウ、ヨウキヒ、イチヨウ等のサトザクラ(Prunus lannesiana)、ソメイヨシノ(Prunus×yedoensis)、カンヒザクラ、エドヒガン、マメザクラ、チョウジザクラ等が挙げられるが、本発明のサクラはこれに限定されるものではない。また、使用部位としては、樹皮、花、種子、実、葉、根等のいずれを用いても良いが、樹皮又は花の使用が好ましい。 In the present invention, "Sakura" is a plant belonging to the genus Plum of the Rosaceae family, for example, Yamazakura (Prunus jamasakura) such as Oshimazakura, Oyamazakura, Kasumizakura, Yamazakura (Yoshino cherry tree), Yaezakura, Kanzan, Fugenzo, Yoshino cherry tree, Examples thereof include Satozakura (Prunus lannesiana) such as Ichiyo, Someiyoshino (Prunus × yedoensis), Kanhizakura, Edohigan, Mamezakura, Choujizakura, etc., but the cherry tree of the present invention is not limited thereto. Further, as the site of use, any of bark, flowers, seeds, fruits, leaves, roots and the like may be used, but the use of bark or flowers is preferable.
サクラの抽出物の調製は、まず、その使用部位を、必要ならば予め水洗して異物を除いた後、そのまま又は乾燥した上、必要に応じて細切又は粉砕し、抽出溶媒と接触させて抽出を行う。抽出は、浸漬法等の常法に従って抽出溶媒と接触させることで行うことが可能であるが、浸漬法以外にも水蒸気蒸留法や超臨界抽出法を用いることも可能である。 To prepare the extract of cherry blossoms, first, the site to be used is washed with water in advance to remove foreign substances, if necessary, and then left as it is or dried, and if necessary, shredded or crushed and brought into contact with an extraction solvent. Extract. Extraction can be carried out by contacting with an extraction solvent according to a conventional method such as a dipping method, but a steam distillation method or a supercritical extraction method can also be used in addition to the dipping method.
抽出溶媒としては、水;メタノール、エタノール、プロパノール等の低級アルコール類;エチレングリコール、1,2−プロパンジオール、1,3−プロパンジオール、1,3−ブチレングリコール、グリセリン等の多価アルコール類;酢酸エチル、酢酸ブチル、プロピオン酸メチル等のエステル類;アセトン、メチルエチルケトン等のケトン類;エチルエーテル、イソプロピルエーテル等のエーテル類;n−ヘキサン、トルエン、クロロホルム等の炭化水素系溶媒等が挙げられ、それらは単独で又は二種以上混合して用いることができる。 As the extraction solvent, water; lower alcohols such as methanol, ethanol and propanol; polyhydric alcohols such as ethylene glycol, 1,2-propanediol, 1,3-propanediol, 1,3-butylene glycol and glycerin; Esters such as ethyl acetate, butyl acetate and methyl propionate; ketones such as acetone and methyl ethyl ketone; ethers such as ethyl ether and isopropyl ether; hydrocarbon solvents such as n-hexane, toluene and chloroform are included. They can be used alone or in admixture of two or more.
抽出溶媒のうちでも、水、低級アルコール類又は多価アルコール類等の親水性溶媒が好適である。この親水性溶媒を用いる場合の好ましい例としては、例えば、水、低級アルコール類(特にエタノール)、又は多価アルコール(特に、1,2−プロパンジオール、1,3−プロパンジオール、1,3−ブチレングリコール)の単独使用、或いは、水と低級アルコール類との混合溶媒、又は水と多価アルコール類との混合溶媒の使用等が挙げられるが、なかでも水単独、或いは水と1,2−プロパンジオール、1,3−プロパンジオール又は1,3−ブチレングリコールの混合溶媒が特に好ましい。 Among the extraction solvents, hydrophilic solvents such as water, lower alcohols and polyhydric alcohols are preferable. Preferred examples of using this hydrophilic solvent include, for example, water, lower alcohols (particularly ethanol), or polyhydric alcohols (particularly 1,2-propanediol, 1,3-propanediol, 1,3-. Butylene glycol) can be used alone, or a mixed solvent of water and lower alcohols, or a mixed solvent of water and polyhydric alcohols can be used. Among them, water alone or water and 1,2- A mixed solvent of propanediol, 1,3-propanediol or 1,3-butylene glycol is particularly preferable.
混合溶媒を用いる場合の混合比は、例えば、水と1,2−プロパンジオール、1,3−プロパンジオール又は1,3−ブチレングリコールとの混合溶媒であれば、容量比(以下同じ)で1:5〜20:1、水とエタノールとの混合溶媒であれば、1:5〜25:1、水とグリセリンとの混合溶媒であれば1:10〜20:1の範囲とすることが好ましい。 When a mixed solvent is used, for example, in the case of a mixed solvent of water and 1,2-propanediol, 1,3-propanediol or 1,3-butylene glycol, the volume ratio (hereinafter the same applies) is 1. : 5 to 20: 1, preferably in the range of 1: 5 to 25: 1 for a mixed solvent of water and ethanol, and preferably in the range of 1: 10 to 20: 1 for a mixed solvent of water and glycerin. ..
また、サクラの使用部位と抽出溶媒との重量比は、好ましくは1:1〜1:100であり、より好ましくは、1:5〜1:60である。 The weight ratio of the site where the cherry tree is used and the extraction solvent is preferably 1: 1 to 1: 100, more preferably 1: 5 to 1:60.
抽出物の調製時のpHに限定はないが、一般には3〜9の範囲とすることが好ましい。かかる意味で、必要であれば、前記抽出溶媒に、水酸化ナトリウム、炭酸ナトリウム、水酸化カリウム等のアルカリ性調整剤、又はクエン酸、塩酸、リン酸、硫酸等の酸性調整剤を配合し、所望のpHとなるように調整してもよい。 The pH at the time of preparation of the extract is not limited, but is generally preferably in the range of 3 to 9. In this sense, if necessary, an alkali adjusting agent such as sodium hydroxide, sodium carbonate, potassium hydroxide or the like, or an acid adjusting agent such as citric acid, hydrochloric acid, phosphoric acid, sulfuric acid, etc. is added to the extraction solvent, which is desired. The pH may be adjusted to the above.
抽出温度、抽出時間等の抽出条件は、用いる溶媒の種類やpHによっても異なるが、例えば、水又は1,3−ブチレングリコール、或いは水と1,2−プロパンジオール、1,3−プロパンジオール又は1,3−ブチレングリコールとの混液を溶媒とする場合であれば、抽出温度は好ましくは0℃〜90℃の範囲であり、又抽出時間は好ましくは1〜168時間(1時間〜1週間)であり、より好ましくは1〜120時間(1時間〜5日間)の範囲である。 Extraction conditions such as extraction temperature and extraction time vary depending on the type of solvent used and pH, but for example, water or 1,3-butylene glycol, or water and 1,2-propanediol, 1,3-propanediol or When a mixed solution with 1,3-butylene glycol is used as a solvent, the extraction temperature is preferably in the range of 0 ° C. to 90 ° C., and the extraction time is preferably 1 to 168 hours (1 hour to 1 week). It is more preferably in the range of 1 to 120 hours (1 hour to 5 days).
なお、本発明の抽出処理に先立って、又は抽出処理と並行して、必要に応じて抽出物に加水分解処理を施してもよい。これによって、抽出物の保存安定性等を改善して、皮膚外用剤や飲食品の機能性素材としての抽出物をより有効に利用できる可能性がある。 If necessary, the extract may be hydrolyzed prior to the extraction treatment of the present invention or in parallel with the extraction treatment. As a result, there is a possibility that the storage stability of the extract can be improved and the extract as a functional material for external preparations for skin and foods and drinks can be used more effectively.
抽出物に酵素加水分解処理を施す場合、酵素としては、アクチナーゼ、パパイン、ペプシン等の蛋白分解酵素、グルコアミラーゼ、α−アミラーゼ、β−アミラーゼ等の澱粉分解酵素、セルラーゼ、ヘミセルラーゼ、ペクチナーゼ等の繊維素分解酵素、リパーゼ等の脂質分解酵素等のいずれかの酵素群から選ばれた1種又は2種以上を用いてもよいが、それらの酵素群からそれぞれ選ばれた1種又は2種以上の酵素を組み合わせて用いることがより好ましい。 When the extract is subjected to enzyme hydrolysis treatment, the enzymes include proteolytic enzymes such as actinase, papaine and pepsin, starch degrading enzymes such as glucoamylase, α-amylase and β-amylase, cellulase, hemicellulase and pectinase. One or more selected from any of the enzyme groups such as fibrinolytic enzyme and lipid degrading enzyme such as lipase may be used, but one or more selected from those enzyme groups, respectively. It is more preferable to use the above enzymes in combination.
酵素の添加量は、例えば、植物の使用部位の固形分に対して、合計で0.00001〜10重量%の範囲とすることが好ましい。 The amount of the enzyme added is preferably in the range of 0.00001 to 10% by weight in total with respect to the solid content of the site where the plant is used.
本発明において、酵母とは、例えば、サッカロミセス セレビシエ(Saccharomyces cerevisiae)、サッカロミセス アワモリ(Saccharomyces awamori)、サッカロミセス チェバリエリ(Saccharomyces chevalieri)、サッカロミセス カールスバージェンシス(Saccharomyces carlsbergensis)、サッカロミセス バヨナス(Saccharomyces bayonus)等のサッカロミセス属の酵母、ガラクトミセス(Galactomyces)属の酵母、トルラスポラ デルブルエキ(Torulaspora delbruekii)、トルラスポラ ファーメンタチ(Torulaspora fermentati)、トルラスポラ ロゼイ(Torulaspora rosei)等のトルラスポラ属の酵母、ジゴサッカロミセス ローキシ(Zygosaccharomyces rouxii)、ジゴサッカロミセス ソーヤ(Zygosacchar omyces soya)、ジゴサッカロミセス サケ(Zygosaccharomyces sake)、ジゴサッカロミセス ミソ(Zygosaccharomyces miso)、ジゴサッカロミセス ラクティス(Zygosaccharomyces lactis)等のジゴサッカロミセス属の酵母、カンディダ ベルサチリス(Candida versatilis)、カンディダ エチェリシイ(Candida etchellsii)、カンディダ ケフィール(Candida kefyr)、カンディダ サケ(Candida sake)、カンディダ スコッティ(Candida scottii)等のカンディダ属の酵母、オーレオバシディウムプルランス(Aureobasidium Pullulans)、オーレオバシディウム マンソニー(Aureobasidium mansonii)、オーレオバシディウム マイクロスティクタム(Aureobasideium microstictum)等のオーレオバシディウム属の酵母などが挙げられる。また、本発明に係る酵母としては、清酒酵母、ワイン酵母、ビール酵母、植物の花(バラ、ユリ、サクラ等)由来の酵母、海由来の酵母の何れであっても良い。 In the present invention, yeast is, for example, Saccharomyces cerevisiae, Saccharomyces awamori, Saccharomyces chevalieri, Saccharomyces cerevisiae, Saccharomyces calsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis, Saccharomyces carlsbergensis. Yeasts of the genus Galactomyces, yeasts of the genus Galactomyces, yeasts of the genus Torulaspora delbruekii, Torulaspora fermentati, Torulaspora rosei, etc. Zygosaccharomyces soya, Zygosaccharomyces sake, Zygosaccharomyces miso, Zygosaccharomyces lactis, Zygosaccharomyces lactis, Zygosaccharomyces lactis, etc. ), Candida kefyr, Candida sake, Candida scottii and other Candida yeasts, Aureobasidium Pullulans, Aureobasidium mansonii , Aureobasideium microstictum and other yeasts of the genus Aureobasideium. Further, the yeast according to the present invention may be any of sake yeast, wine yeast, brewer's yeast, yeast derived from plant flowers (rose, lily, cherry tree, etc.), and yeast derived from the sea.
本発明において、麹菌としては、例えばアスペルギルス オリゼー(Aspergillus oryzae)、アスペルギルス フラバス(Aspergillus flavus)、アスペルギルス ポリオキソジェネス(Aspergillus polyoxogenes)、アスペルギルス ソーヤ(Aspergillus sojae)等の黄麹菌、アスペルギルス アワモリ(Aspergillus awamori)、アスペルギルス カワウチ(Aspergillus kawauchii)、アスペルギルス ウサミ(Aspergillus usami)、アスペルギルス ニガー(Aspergillus niger)等の黒麹菌、モナスカス アンカ(Monascus anka)、モナスカス ピロサス(Monascus pilosus)等の紅麹菌などが挙げられる。 In the present invention, examples of aspergillus include Aspergillus oryzae, Aspergillus flavus, Aspergillus polyoxogenes, Aspergillus sojae, Aspergillus aspergillus, Aspergillus aspergillus, and Aspergillus aspergillus. Aspergillus kawauchii, Aspergillus usami, Aspergillus niger and other black aspergillus, Monascus anka, Monascus pilosus and other red aspergillus.
本発明において、乳酸菌とは、例えばラクトバシルス プランタラム(Lactobacillus plantarum)、ラクトバシルス ブレビス(L. brevis)、ラクトバシルス カゼイ(Lactobacillus casei)、ラクトバチルス・デルブルッキー(Lactobacillus delbrueckii)等のラクトバシルス(Lactobacillus)属の乳酸菌;カルノバクテリウム ディバージェンス(Carnobacterium divergens)、カルノバクテリウム ピシコーラ(Carnobacterium piscicola)等のカルノバクテリウム(Carnobacterium)属の乳酸菌;ロイコノストック メセンテロイズ(Leuconostoc mesenteroides)、ロイコノストック シトレウム(Leuconostoc citreum)等のロイコノストック(Leuconostoc)属の乳酸菌; ストレプトコッカス フェーカリス(Streptococcus faecalis)、ストレプトコッカス ピオジェネス(Streptococcus pyogenes)等のストレプトコッカス属の乳酸菌;エンテロコッカス カゼリフラバス(Enterococcus caseliflavus)、エンテロコッカス サルフレウス(Enterococcus sulfreus)等のエンテロコッカス(Enterococcus)属の乳酸菌;ラクトコッカス プランタラム(Lactococcus plantarum) ラクトコッカス ラフィノラクティス(Lactococcus rafinolactis)等のラクトコッカス属の乳酸菌;ヴェイセラ コンフューザ(Weissella confusa)、ヴェイセラ カンドウレリ(Weissella kandleri)等のヴェイセラ属の乳酸菌;アトポビウム ミニュタム(Atopobium minutum)、アトポビウム パービュラス(Atopobiumparvulus)等のアトポビウム(Atopobium)属の乳酸菌;バゴコッカス フルビアリス(Vagococcus fluvialis)、バゴコッカス サーモニナラム(Vagococcus salmoninarum)等のバゴコッカス(Vagococcus)属の乳酸菌;ペディオコッカス ダムノサス(Pediococcus damnosus)、ペディオコッカス ペントサセウス(Pediococcus pentosaceus)等のペディオコッカス(Pediococcus)属の乳酸菌等が挙げられる。 In the present invention, the lactic acid bacterium is, for example, a lactic acid bacterium of the genus Lactobacillus delbrueckii such as Lactobacillus plantarum, L. brevis, Lactobacillus casei, and Lactobacillus delbrueckii. Lactic acid bacteria of the genus Carnobacterium such as Carnobacterium divergens and Carnobacterium piscicola; Leuconostoc mesenteroides, Leuconostoc citreum, etc. Lactic acid bacteria of the genus Leuconostoc; Lactic acid bacteria of the genus Streptococcus such as Streptococcus faecalis and Streptococcus pyogenes; Lactic acid bacteria of the genus; Lactococcus plantarum Lactococcus rafinolactis and other lactic acid bacteria of the genus Lactococcus; Weissella confusa, Weissella kandleri and other lactic acid bacteria of the genus Weissella kandleri (Atopobium minutum), Atopobium parvulus and other lactic acid bacteria of the genus Atopobium; Vagococcus fluvialis, Vagococcus fluvialis, Vagococcus salmoninarum, etc. Examples thereof include lactic acid bacteria belonging to the genus Pediococcus such as Su (Pediococcus damnosus) and Pediococcus pentosaceus.
本発明において、酵母、麹菌又は乳酸菌を培地で培養し、濾過等の操作により培養液を回収することで、微生物の抽出物を得ることができる。また、それら微生物のいずれか1種以上を用いて米、ハトムギ、大豆等の植物を発酵させて得られる発酵物を微生物由来発酵代謝物として、本発明に係る組成物の有効成分としても良い。この場合、微生物由来発酵代謝物として、発酵物中の固形分相を用いても、又は発酵物から固形分を除いて得られる発酵液を用いても良い。 In the present invention, an extract of a microorganism can be obtained by culturing yeast, aspergillus or lactic acid bacterium in a medium and collecting the culture solution by an operation such as filtration. In addition, a fermented product obtained by fermenting a plant such as rice, adlay, soybean, etc. using any one or more of these microorganisms may be used as a fermentation metabolite derived from a microorganism as an active ingredient of the composition according to the present invention. In this case, as the microbial-derived fermentation metabolite, the solid content phase in the fermented product may be used, or a fermented liquid obtained by removing the solid content from the fermented product may be used.
また、本発明においては、微生物由来発酵代謝物として、酵母、麹菌及び/又は乳酸菌に用いて製造される醸造酒又は蒸留酒、或いは醸造酒又は蒸留酒の製造過程で生じる残渣(酒粕、焼酎粕、ワイン粕、ビール粕)及びそれら残渣から抽出処理により得られる抽出物を用いることもできる。 Further, in the present invention, as a fermentation metabolite derived from microorganisms, brewed liquor or distilled liquor produced by using yeast, aspergillus and / or lactic acid bacterium, or residue (sake lees, liquor lees) produced in the production process of brewed liquor or distilled liquor. , Wine lees, beer lees) and extracts obtained by extraction treatment from their residues can also be used.
本発明の組成物は、例えば、皮膚(頭皮も含む)外用剤(化粧料、医薬部外品、外用医薬品)、美容用又は健康増進用の飲食品に配合することができる。皮膚外用剤としては、例えば、乳液、クリーム、ローション、エッセンス、パック、口紅、ファンデーション、リクイドファンデーション、メイクアッププレスパウダー、ほほ紅、白粉、洗顔料、ボディシャンプー、頭皮,頭髪用シャンプー、頭髪用コンディショナー、育毛,養毛用のシャンプー又はトニック、石けん等の清浄用化粧料、さらには浴剤等が挙げられるが、本発明はこれらに限定されるものではない。また、美容用又は健康増進用の飲食品としては、美容飲料、栄養ドリンク、スポーツドリンク、ニアウォーター、ビタミン飲料、ミネラル飲料、アルコール飲料等の飲料;各種スープ類(粉末スープも含む)、乳製品、ゼリー、キャンディ、錠菓、ガム等の食品;錠剤、液状、顆粒状又はゼリー状の健康食品・飲料等に配合することができるが、本発明はこれらに限るものではなく、経口摂取できる飲食品等に配合することができる The composition of the present invention can be incorporated into, for example, skin (including scalp) external preparations (cosmetics, quasi-drugs, external medicines), and foods and drinks for beauty or health promotion. Examples of external skin preparations include emulsions, creams, lotions, essences, packs, lipsticks, foundations, liquid foundations, make-up press powders, blushers, white powder, pigment wash, body shampoos, scalp, hair shampoos, and hair conditioners. , Shampoo or tonic for hair growth and hair growth, cleansing cosmetics such as soap, and bathing agents, but the present invention is not limited thereto. In addition, as foods and drinks for beauty or health promotion, beverages such as beauty beverages, energy drinks, sports drinks, near water, vitamin beverages, mineral beverages, alcoholic beverages; various soups (including powdered soups), dairy products , Jelly, candy, tablet confectionery, gum, etc .; Although it can be blended in tablets, liquid, granular or jelly-like health foods and beverages, the present invention is not limited to these, and foods and drinks that can be taken orally. Can be blended in products, etc.
本発明の組成物の配合量は、組成物の固形分として、基礎化粧料の場合は、0.002〜1.0重量%(固形分重量%、以下同じ)の範囲、メイクアップ化粧料の場合は、0.002〜1.0重量%の範囲、又清浄用化粧料の場合は、0.002〜10.0重量%の範囲である。また、毛髪用化粧料の場合は、組成物の固形分として、0.0001〜5.0重量%の範囲である。また、飲食品おける本発明の組成物の配合量は、組成物の固形分として、0.1〜15重量%の範囲が好ましい。 The blending amount of the composition of the present invention shall be in the range of 0.002 to 1.0% by weight (solid content weight%, the same shall apply hereinafter) in the case of basic cosmetics as the solid content of the composition, and of makeup cosmetics. In the case, it is in the range of 0.002 to 1.0% by weight, and in the case of cosmetics for cleaning, it is in the range of 0.002 to 10.0% by weight. In the case of hair cosmetics, the solid content of the composition is in the range of 0.0001 to 5.0% by weight. The blending amount of the composition of the present invention in foods and drinks is preferably in the range of 0.1 to 15% by weight as the solid content of the composition.
本発明の組成物を皮膚外用剤又は飲食品に配合する場合、必須成分である組成物のほかに、通常のそれら製品に用いられる成分、例えば油性成分、界面活性剤(合成系、天然物系)、保湿剤、増粘剤、防腐・殺菌剤、粉体成分、紫外線吸収剤、抗酸化剤、色素、香料等を必要に応じて適宜配合することができる。また、当該組成物の有効性、特長を損なわない限り、他の生理活性成分を組み合わせて配合することも何ら差し支えない。 When the composition of the present invention is blended with an external preparation for skin or food and drink, in addition to the composition which is an essential component, components usually used in those products, such as an oil component and a surfactant (synthetic system, natural product system) ), Moisturizer, thickener, preservative / bactericidal agent, powder component, ultraviolet absorber, antioxidant, pigment, fragrance and the like can be appropriately blended as needed. Further, as long as the effectiveness and features of the composition are not impaired, other physiologically active ingredients may be combined and blended at all.
油性成分としては、例えばハス油、オリーブ油、ホホバ油、ヒマシ油、大豆油、米油、米糠油、米胚芽油、ヤシ油、カミツレ油、パーム油、カカオ油、メドウフォーム油、ローズヒップ油、バラ油、ランベンダー油、シアーバター、ティーツリー油、アボガド油、マカデミアナッツ油、ベルガモット油、植物由来スクワラン等の植物由来の油脂類;ミンク油、タートル油等の動物由来の油脂類;ミツロウ、カルナウバロウ、ライスワックス、ラノリン等のロウ類;流動パラフィン、ワセリン、パラフィンワックス、スクワラン等の炭化水素類;ミリスチン酸、パルミチン酸、ステアリン酸、オレイン酸、イソステアリン酸、エイコセン酸等の脂肪酸類;ラウリルアルコール、セタノール、ステアリルアルコール等の高級アルコール類;ミリスチン酸イソプロピル、パルミチン酸イソプロピル、オレイン酸ブチル、2−エチルヘキシルグリセライド、高級脂肪酸オクチルドデシル(ステアリン酸オクチルドデシル等)等の合成エステル類及び合成トリグリセライド類等が挙げられる。 Oily components include, for example, lotus oil, olive oil, jojoba oil, castor oil, soybean oil, rice oil, rice bran oil, rice germ oil, palm oil, chamomile oil, palm oil, cacao oil, meadowfoam oil, rosehip oil, etc. Rose oil, lanbender oil, sheer butter, tea tree oil, avocado oil, macadamia nut oil, bergamot oil, plant-derived oils and fats such as squalane; animal-derived oils and fats such as mink oil and turtle oil; , Rice wax, waxes such as lanolin; hydrocarbons such as liquid paraffin, vaseline, paraffin wax, squalane; fatty acids such as myristic acid, palmitic acid, stearic acid, oleic acid, isostearic acid, eicosenoic acid; lauryl alcohol, Higher alcohols such as cetanol and stearyl alcohol; synthetic esters such as isopropyl myristate, isopropyl palmitate, butyl oleate, 2-ethylhexyl glyceride, higher fatty acid octyldodecyl (octyldodecyl stearate, etc.) and synthetic triglycerides, etc. Be done.
界面活性剤としては、例えばポリオキシエチレンアルキルエーテル、ポリオキシエチレン脂肪酸エステル、ポリオキシエチレンソルビタン脂肪酸エステル、グリセリン脂肪酸エステル、ポリグリセリン脂肪酸エステル、ポリオキシエチレングリセリン脂肪酸エステル、ポリオキシエチレン硬化ヒマシ油、ポリオキシエチレンソルビトール脂肪酸エステル等の非イオン界面活性剤;脂肪酸塩、アルキル硫酸塩、アルキルベンゼンスルホン酸塩、ポリオキシエチレンアルキルエーテル硫酸塩、ポリオキシエチレン脂肪アミン硫酸塩、ポリオキシエチレンアルキルフェニルエーテル硫酸塩、ポリオキシエチレンアルキルエーテル燐酸塩、α−スルホン化脂肪酸アルキルエステル塩、ポリオキシエチレンアルキルフェニルエーテル燐酸塩等のアニオン界面活性剤;第四級アンモニウム塩、第一級〜第三級脂肪アミン塩、トリアルキルベンジルアンモニウム塩、アルキルピリジニウム塩、2−アルキル−1−アルキル−1−ヒドロキシエチルイミダゾリニウム塩、N,N−ジアルキルモルフォルニウム塩、ポリエチレンポリアミン脂肪酸アミド塩等のカチオン界面活性剤;N,N−ジメチル−N−アルキル−N−カルボキシメチルアンモニオベタイン、N,N,N−トリアルキル−N−アルキレンアンモニオカルボキシベタイン、N−アシルアミドプロピル−N′,N′−ジメチル−N′−β−ヒドロキシプロピルアンモニオスルホベタイン等の両性界面活性剤等を使用することができる。 Examples of the surfactant include polyoxyethylene alkyl ether, polyoxyethylene fatty acid ester, polyoxyethylene sorbitan fatty acid ester, glycerin fatty acid ester, polyglycerin fatty acid ester, polyoxyethylene glycerin fatty acid ester, polyoxyethylene hydrogenated castor oil, and poly. Nonionic surfactants such as oxyethylene sorbitol fatty acid esters; fatty acid salts, alkyl sulfates, alkylbenzene sulfonates, polyoxyethylene alkyl ether sulfates, polyoxyethylene fatty amine sulfates, polyoxyethylene alkylphenyl ether sulfates, Anionic surfactants such as polyoxyethylene alkyl ether phosphates, α-sulfonated fatty acid alkyl ester salts, polyoxyethylene alkyl phenyl ether phosphates; quaternary ammonium salts, primary to tertiary fatty amine salts, tri Cationic surfactants such as alkylbenzylammonium salt, alkylpyridinium salt, 2-alkyl-1-alkyl-1-hydroxyethylimidazolinium salt, N, N-dialkylmorphonium salt, polyethylene polyamine fatty acid amide salt; N, N-dimethyl-N-alkyl-N-carboxymethylammoniobetaine, N, N, N-trialkyl-N-alkylene ammoniocarboxybetaine, N-acylamide propyl-N', N'-dimethyl-N'- An amphoteric surfactant such as β-hydroxypropylammoniosulfobetaine can be used.
乳化剤又は乳化助剤としては、酵素処理ステビア等のステビア誘導体、サポニン又はその誘導体、カゼイン又はその塩(ナトリウム等)、糖と蛋白質の複合体、ショ糖又はそのエステル、ラクトース、大豆由来の水溶性多糖、大豆由来蛋白質と多糖の複合体、ラノリン又はその誘導体、コレステロール、ステビア誘導体(ステビア酵素処理物等)、ケイ酸塩(アルミニウム、マグネシウム等)、炭酸塩(カルシウム、ナトリウム等)、サポニン及びその誘導体、レシチン及びその誘導体(水素添加レシチン等)、乳酸菌醗酵米、乳酸菌醗酵発芽米、乳酸菌醗酵穀類(麦類、豆類、雑穀等)等を配合することもできる。 Examples of emulsifiers or emulsifying aids include stevia derivatives such as enzyme-treated stevia, saponins or derivatives thereof, casein or salts thereof (sodium, etc.), sugar-protein complexes, sucrose or esters thereof, lactose, and water-soluble soybeans. Polysaccharide, complex of soybean-derived protein and polysaccharide, lanolin or its derivative, cholesterol, stevia derivative (stevia enzyme treated product, etc.), silicate (aluminum, magnesium, etc.), carbonate (calcium, sodium, etc.), saponin and its Derivatives, lecithin and its derivatives (hydrogenated lecithin, etc.), lactic acid bacteria fermented rice, lactic acid bacteria fermented sprouted rice, lactic acid bacteria fermented grains (wheat, beans, miscellaneous grains, etc.) and the like can also be blended.
保湿剤としては、例えばグリセリン、プロピレングリコール、ジプロピレングリコール、1,3−ブチレングリコール、ポリエチレングリコール、ソルビトール、キシリトール、ピロリドンカルボン酸ナトリウム等があり、さらにトレハロース等の糖類、ムコ多糖類(例えば、ヒアルロン酸及びその誘導体、コンドロイチン及びその誘導体、ヘパリン及びその誘導体等)、エラスチン及びその誘導体、コラーゲン及びその誘導体、NMF関連物質、乳酸、尿素、高級脂肪酸オクチルドデシル、海藻抽出物、シラン根(白及)抽出物、各種アミノ酸及びそれらの誘導体が挙げられる。 Examples of the moisturizing agent include glycerin, propylene glycol, dipropylene glycol, 1,3-butylene glycol, polyethylene glycol, sorbitol, xylitol, sodium pyrrolidone carboxylate and the like, and saccharides such as trehalose and mucopolysaccharides (for example, hyalurone). Acids and their derivatives, chondroitin and its derivatives, heparin and its derivatives, etc.), elastin and its derivatives, collagen and its derivatives, NMF-related substances, lactic acid, urea, higher fatty acids octyldodecyl, seaweed extract, silane roots (white) Examples include extracts, various amino acids and their derivatives.
増粘剤としては、例えばアルギン酸、寒天、カラギーナン、フコイダン等の褐藻、緑藻又は紅藻由来成分;シラン根(白及)抽出物;ペクチン、ローカストビーンガム、アロエ多糖体、アルカリゲネス産生多糖体等の多糖類;キサンタンガム、トラガントガム、グアーガム等のガム類;カルボキシメチルセルロース、ヒドロキシエチルセルロース等のセルロース誘導体;ポリビニルアルコール、ポリビニルピロリドン、カルボキシビニルポリマー、アクリル酸・メタクリル酸共重合体等の合成高分子類;ヒアルロン酸及びその誘導体;ポリグルタミン酸及びその誘導体等が挙げられる。 Examples of the thickener include brown algae such as alginic acid, agar, carrageenan, and fucoidan, components derived from green algae or red algae; silane root (white and) extract; pectin, locust bean gum, aloe polysaccharide, alkaligenes-producing polysaccharide, and the like. Polysaccharides; gums such as xanthan gum, tragant gum, guar gum; cellulose derivatives such as carboxymethyl cellulose and hydroxyethyl cellulose; synthetic polymers such as polyvinyl alcohol, polyvinylpyrrolidone, carboxyvinyl polymer, alginic acid / methacrylic acid copolymer; hyaluronic acid And derivatives thereof; examples thereof include polyglutamic acid and derivatives thereof.
防腐・殺菌剤としては、例えば尿素;パラオキシ安息香酸メチル、パラオキシ安息香酸エチル、パラオキシ安息香酸プロピル、パラオキシ安息香酸ブチル等のパラオキシ安息香酸エステル類;フェノキシエタノール、ジクロロフェン、ヘキサクロロフェン、塩酸クロルヘキシジン、塩化ベンザルコニウム、サリチル酸、エタノール、ウンデシレン酸、フェノール類、ジャマール(イミダゾデイニールウレア)、ポリリン酸、プロパンジオール、1,2−ペンタンジオール、各種精油類、樹皮乾留物、大根発酵液、サトウキビ等の植物由来のエタノール又は1,3−ブチレングリコール等がある。 Examples of antiseptic / bactericidal agents include urea; paraoxybenzoic acid esters such as methyl paraoxybenzoate, ethyl paraoxybenzoate, propyl paraoxybenzoate, and butyl paraoxybenzoate; phenoxyethanol, dichlorophene, hexachlorophene, chlorhexidine hydrochloride, and benza chloride. Plants such as luconium, salicylic acid, ethanol, undesyleneic acid, phenols, jamar (imidazodeine luurea), polyphosphate, propanediol, 1,2-pentanediol, various essential oils, dried bark, radish fermented liquid, sugar cane, etc. Derived ethanol or 1,3-butylene glycol and the like.
粉体成分としては、例えばセリサイト、酸化チタン、タルク、カオリン、ベントナイト、酸化亜鉛、炭酸マグネシウム、酸化マグネシウム、酸化ジルコニウム、硫酸バリウム、無水ケイ酸、雲母、ナイロンパウダー、ポリエチレンパウダー、シルクパウダー、セルロース系パウダー、穀類(米、麦、トウモロコシ、キビ等)のパウダー、豆類(大豆、小豆等)のパウダー等がある。 Examples of powder components include cericite, titanium oxide, talc, kaolin, bentonite, zinc oxide, magnesium carbonate, magnesium oxide, zirconium oxide, barium sulfate, silicic anhydride, mica, nylon powder, polyethylene powder, silk powder, and cellulose. There are system powders, grain powders (rice, wheat, corn, talc, etc.), beans (soybeans, talc, etc.) powders, etc.
紫外線吸収剤としては、例えばパラアミノ安息香酸エチル、パラジメチルアミノ安息香酸エチルヘキシル、サリチル酸アミル及びその誘導体、パラメトキシ桂皮酸2−エチルヘキシル、桂皮酸オクチル、オキシベンゾン、2,4−ジヒドロキシベンゾフェノン、2−ヒドロキシ−4−メトキシベンゾフェノン−5−スルホン酸塩、4−ターシャリーブチル−4−メトキシベンゾイルメタン、2−(2−ヒドロキシ−5−メチルフェニル)ベンゾトリアゾール、ウロカニン酸、ウロカニン酸エチル、アロエ抽出物等がある。 Examples of the ultraviolet absorber include ethyl paraaminobenzoate, ethylhexyl paradimethylaminobenzoate, amyl salicylate and its derivatives, 2-ethylhexyl paramethoxycinnamate, octyl silicate, oxybenzone, 2,4-dihydroxybenzophenone, 2-hydroxy-4. -Methoxybenzophenone-5-sulfonate, 4-tershally butyl-4-methoxybenzoylmethane, 2- (2-hydroxy-5-methylphenyl) benzotriazole, urocanic acid, ethyl urocanate, aloe extract, etc. ..
抗酸化剤としては、例えばブチルヒドロキシアニソール、ブチルヒドロキシトルエン、没食子酸プロピル、ムラサキシキブの抽出物、シラン根の抽出物、シャクヤク抽出物、ビタミンE及びその誘導体(例えば、ビタミンEニコチネート、ビタミンEリノレート等)等がある。 Antioxidants include, for example, butylhydroxyanisole, butylhydroxytoluene, propyl gallate, beautyberry extract, silane root extract, shakuyaku extract, vitamin E and its derivatives (for example, vitamin E nicotinate, vitamin E linoleate, etc.) ) Etc.
美白剤としては、t−シクロアミノ酸誘導体、コウジ酸及びその誘導体、アスコルビン酸及びその誘導体、ハイドロキノン又はその誘導体、エラグ酸及びその誘導体、ニコチン酸及びその誘導体、レゾルシノール誘導体、トラネキサム酸及びその誘導体、4−メトキシサリチル酸カリウム塩、マグノリグナン(5,5'−ジプロピル−ビフェニル−2,2’−ジオール)、4−(4−ヒドロキシフェニル)−4−ブタノール))、ヒドロキシ安息香酸及びその誘導体、ビタミンE及びその誘導体、α−ヒドロキシ酸、AMP(アデノシンモノホスフェイト、アデノシン1リン酸)が挙げられ、これらを単独で配合しても、複数を組み合わせて配合しても良い。 Whitening agents include t-cycloamino acid derivatives, kodiic acid and its derivatives, ascorbic acid and its derivatives, hydroquinone or its derivatives, ellagic acid and its derivatives, nicotinic acid and its derivatives, resorcinol derivatives, tranexamic acid and its derivatives, 4 -Potasium salicylic acid salt, magnolignan (5,5'-dipropyl-biphenyl-2,2'-diol), 4- (4-hydroxyphenyl) -4-butanol)), hydroxybenzoic acid and its derivatives, vitamin E And derivatives thereof, α-hydroxy acid, and AMP (adenosine monophosphate, adenosine monophosphate), and these may be blended alone or in combination of two or more.
上記のコウジ酸誘導体としては、例えばコウジ酸モノブチレート、コウジ酸モノカプレート、コウジ酸モノパルミテート、コウジ酸ジブチレート等のコウジ酸エステル類、コウジ酸エーテル類、コウジ酸グルコシド等のコウジ酸糖誘導体等が、アスコルビン酸誘導体としては、例えばL−アスコルビン酸−2−リン酸エステルナトリウム、L−アスコルビン酸−2−リン酸エステルマグネシウム、L−アスコルビン酸−2−硫酸エステルナトリウム、L−アスコルビン酸−2−硫酸エステルマグネシウム等のアスコルビン酸エステル塩類、L−アスコルビン酸−2−グルコシド、L−アスコルビン酸−5−グルコシド等のアスコルビン酸糖誘導体、それらアスコルビン酸糖誘導体の6位アシル化物(アシル基は、ヘキサノイル基、オクタノイル基、デカノイル基等)、L−アスコルビン酸テトライソパルミチン酸エステル、L−アスコルビン酸テトララウリン酸エステル等のL−アスコルビン酸テトラ脂肪酸エステル類、3−O−エチルアスコルビン酸、L−アスコルビン酸−2−リン酸−6−O−パルミテートナトリウム、グリセリルアスコルビン酸又はそのアシル化誘導体、ビスグリセリルアスコルビン酸等のアスコルビン酸グルセリン誘導体が、ハイドロキノン誘導体としては、アルブチン(ハイドロキノン−β−D−グルコピラノシド)、α−アルブチン(ハイドロキノン−α−D−グルコピラノシド)等が、トラネキサム酸誘導体としては、トラネキサム酸エステル(例えば、トラネキサム酸ラウリルエステル、トラネキサム酸ヘキサデシルエステル、トラネキサム酸セチルエステル又はその塩)、トラネキサム酸のアミド体(例えば、トラネキサム酸メチルアミド)等が挙げられ、レゾルシノール誘導体としては、例えば、4−n−ブチルレゾルシノール、4−イソアミルレゾルシノール等が、2,5−ジヒドロキシ安息香酸誘導体としては、例えば2,5−ジアセトキシ安息香酸、2−アセトキシ−5−ヒドロキシ安息香酸、2−ヒドロキシ−5−プロピオニルオキシ安息香酸等が、ニコチン酸誘導体としては、例えばニコチン酸アミド、ニコチン酸ベンジル等が、α−ヒドロキシ酸としては、例えば乳酸、リンゴ酸、コハク酸、クエン酸、α−ヒドロキシオクタン酸等がある。 Examples of the ascorbic acid derivative include ascorbic acid esters such as ascorbic acid monobutyrate, ascorbic acid monocaplate, ascorbic acid monopalmitate, and ascorbic acid dibutyrate, ascorbic acid ethers, and ascorbic acid sugar derivatives such as ascorbic acid glucoside. However, examples of the ascorbic acid derivative include L-ascorbic acid-2-phosphate ester sodium, L-ascorbic acid-2-phosphate ester magnesium, L-ascorbic acid-2-sulfate sodium ester, and L-ascorbic acid-2. -Ascorbic acid ester salts such as magnesium sulfate, ascorbic acid sugar derivatives such as L-ascorbic acid-2-glucoside, L-ascorbic acid-5-glucoside, and 6-position acylated products of these ascorbic acid sugar derivatives (the acyl group is Hexanoyl group, octanoyl group, decanoyl group, etc.), L-ascorbic acid tetrafatty acid esters such as L-ascorbic acid tetraisopalmitic acid ester, L-ascorbic acid tetralauric acid ester, 3-O-ethylascorbic acid, L- Glucerin ascorbic acid derivatives such as ascorbic acid-2-phosphate-6-O-palmitate sodium, glyceryl ascorbic acid or an acylated derivative thereof, and bisglyceryl ascorbic acid are used as hydroquinone derivatives. Glucopyranoside), α-arbutin (hydroquinone-α-D-glucopyranoside), etc., as the tranexamic acid derivative, tranexamic acid ester (for example, tranexamic acid lauryl ester, tranexamic acid hexadecyl ester, tranexamic acid cetyl ester or a salt thereof), Examples thereof include an amide form of tranexamic acid (for example, methylamide tranexamic acid), and examples of the resorcinol derivative include 4-n-butylresorbicinol and 4-isoamylresorbic acid, and examples of the 2,5-dihydroxybenzoic acid derivative include. 2,5-diacetoxybenzoic acid, 2-acetoxy-5-hydroxybenzoic acid, 2-hydroxy-5-propionyloxybenzoic acid and the like, and nicotinic acid derivatives such as nicotinic acid amide and benzyl nicotinate are α- Examples of the hydroxy acid include lactic acid, ascorbic acid, succinic acid, citric acid, α-hydroxyoctanoic acid and the like.
生理活性成分としては、例えば、胎盤抽出液、ソウハクヒ抽出物、ユキノシタ抽出物、シソ抽出物、米糠抽出物又はその加水分解物、白芥子抽出物又はその加水分解物、白芥子の発酵物、シャクヤク抽出物又はその加水分解物、ムラサキシキブ抽出物、ハス種子抽出物又はその加水分解物、ハス種子発酵物、党参抽出物又はその加水分解物、ハトムギ加水分解物、ローヤルゼリー発酵物、パンダヌス・アマリリフォリウス(Pandanus amaryllifolius Roxb.)抽出物、アルカンジェリシア・フラバ(Arcangelicia flava Merrilli)抽出物、カミツレ抽出物等が挙げられる。また、サンゴ草抽出物、イネの葉の抽出物又はその加水分解物、ナス(ベルガモット、長ナス、賀茂ナス、米ナス等)抽出物又はその加水分解物、アンズ果実の抽出物、カタメンキリンサイ等の海藻の抽出物、アマモ等の海産顕花植物の抽出物、豆乳発酵物、クラゲ水、米抽出物又はその加水分解物、発芽米抽出物又はその加水分解物、黒豆抽出物又はその加水分解物、ダマスクバラの花の抽出物、タケノコの皮の抽出物、リノール酸及びその誘導体もしくは加工物(例えばリポソーム化リノール酸等)、動物又は魚由来のコラーゲン及びその誘導体、エラスチン及びその誘導体、グリチルリチン酸及びその誘導体(ジカリウム塩等)、t−シクロアミノ酸誘導体、ビタミンA及びその誘導体、アラントイン、ジイソプロピルアミンジクロロアセテート、γ−アミノ−β−ヒドロキシ酪酸、ゲンチアナ抽出物、甘草抽出物、ニンジン抽出物、オタネニンジン抽出物又はその発酵物、紅参抽出物、ミツイシコンブ抽出物、ヘチマ抽出物、アナアオサ抽出物、モモ抽出物、桃仁抽出物、キウイ抽出物、ヒマワリ抽出物、ジュアゼイロ(Zizyphus joazeiro)抽出物、パウダルコ樹皮抽出物、萱草(デイリリー)抽出物又は発酵物、ハイビスカスの花抽出物又は発酵物、ハゴロモグサ抽出物、チェリモヤ抽出物、マンゴー抽出物、マンゴスチン抽出物、フノリ抽出物、烏龍茶抽出物、紅富貴抽出物、紫蘭抽出物、山椒果皮又は種皮の抽出物又は加水分解物、ベニバナ花抽出物、カサブランカ抽出物、甘藷抽出物又はその発酵物、グアバ葉抽出物、ドクダミ抽出物、晩白柚抽出物、アロエ抽出物、イチジク花抽出物、リンゴ抽出物、ホワイトアスパラガス抽出物等がある。 Physiologically active ingredients include, for example, placenta extract, sohakuhi extract, yukinoshita extract, perilla extract, rice bran extract or its hydrolyzate, white potato extract or its hydrolyzate, white potato fermented product, shakuyaku. Extract or its hydrolyzate, Murasakikib extract, Hass seed extract or its hydrolyzate, Hass seed fermented product, Ginseng extract or its hydrolyzate, Hatomugi hydrolyzate, Royal jelly fermented product, Pandanus amalifoliaus Examples include (Pandanus amaryllifolius Roxb.) Extract, Arcangelicia flava Merrilli extract, chamomile extract and the like. In addition, coral grass extract, rice leaf extract or its hydrolyzate, eggplant (bergamot, long eggplant, Kamo eggplant, rice eggplant, etc.) extract or its hydrolyzate, apricot fruit extract, catamen giraffe, etc. Seaweed extract, extract of marine flowering plants such as Amamo, fermented soymilk, jellyfish water, rice extract or its hydrolyzate, sprouted rice extract or its hydrolyzate, black bean extract or its hydrolysis Extracts of damask rose flowers, extracts of bamboo shoots, linoleic acid and its derivatives or processed products (eg, liposome-derived linoleic acid, etc.), animal or fish-derived collagen and its derivatives, elastin and its derivatives, glycyrrhizin Acids and their derivatives (dipotassium salt, etc.), t-cycloaminomino acid derivatives, vitamin A and its derivatives, allantin, diisopropylamine dichloroacetate, γ-amino-β-hydroxybutyric acid, gentian extract, licorice extract, carrot extract, Otaneninjin extract or its fermented product, Benin extract, Mitsuishikonbu extract, Hechima extract, Anaaosa extract, Peach extract, Peach seed extract, Kiwi extract, Sunflower extract, Zizyphus joazeiro extract, Paudalco Bark extract, daily grass extract or fermented product, hibiscus flower extract or fermented product, hagoromogusa extract, cherimoya extract, mango extract, mangostin extract, funori extract, crow dragon tea extract, Benito Takashi extract , Purple orchid extract, Sansho peel or seed coat extract or hydrolyzate, Benibana flower extract, Casablanca extract, Sweet potato extract or its fermented product, Guava leaf extract, Dokudami extract, Late white yuzu extract, There are aloe extract, fig flower extract, apple extract, white asparagus extract and the like.
次に、製造例、処方例及び試験例によって本発明をさらに具体的に説明するが、本発明はそれらに限定されるものではない。なお、以下において、部はすべて重量部を、また%はすべて重量%を意味する。 Next, the present invention will be described in more detail with reference to Production Examples, Formulation Examples, and Test Examples, but the present invention is not limited thereto. In the following, all parts mean parts by weight, and% means all parts by weight.
製造例1.組成物(1)の調製
(i)サクラの抽出物の調製
サトザクラ(Prunus lannesiana)の花を乾燥、粉砕し、粉砕物18gに精製水900gを加え、80℃にて、2時間抽出後、濾過し、褐色透明のサクラ抽出物溶液1385gを得た(固形分濃度0.49%)。
(ii)微生物由来発酵代謝物
植物(サクラ)の花由来の酵母を、グルコース・ペプトンを含む溶液で、15℃で48時間培養し、培養液を濾過して得られる培養液を酵母抽出物溶液(固形分濃度0.51%)とした。
(iii)組成物の調製
サクラ抽出物溶液と酵母抽出物溶液を、1:1の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.40%)
Production example 1. Preparation of composition (1) (i) Preparation of cherry extract The flowers of Satozakura (Prunus lannesiana) are dried and crushed, 900 g of purified water is added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 2 hours and then filtered. Then, 1385 g of a brown transparent cherry extract solution was obtained (solid content concentration 0.49%).
(Ii) Microorganism-derived fermented metabolite Yeast derived from plant (Sakura) flowers is cultured in a solution containing glucose and peptone at 15 ° C. for 48 hours, and the culture solution obtained by filtering the culture solution is a yeast extract solution. (Solid content concentration 0.51%).
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 40%)
製造例2.組成物(2)の調製
(i)サクラの抽出物の調製
サトザクラの花を乾燥、粉砕し、粉砕物18gに精製水450gと1,3−ブチレングリコール450gを加え、80℃にて2時間抽出後、濾過し、褐色透明のサクラ抽出物溶液1365gを得た(固形分濃度0.45%)。
(ii)微生物由来発酵代謝物の調製
精製水800gに、清酒の製造過程で生じる酒粕160gを加え、80℃にて1時間抽出後、濾過し、淡黄色透明の酒粕抽出物溶液2490gを得た(固形濃度0.45%)
(iii)組成物の調製
サクラの抽出物と酒粕抽出物を、1:9の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.34%)。
Production example 2. Preparation of composition (2) (i) Preparation of cherry extract The flowers of Satozakura are dried and crushed, 450 g of purified water and 450 g of 1,3-butylene glycol are added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 2 hours. Then, it was filtered to obtain 1365 g of a brown transparent cherry extract solution (solid content concentration 0.45%).
(Ii) Preparation of fermented metabolites derived from microorganisms 160 g of sake lees produced in the process of producing sake was added to 800 g of purified water, extracted at 80 ° C. for 1 hour, and then filtered to obtain 2490 g of a pale yellow transparent sake lees extract solution. (Solid concentration 0.45%)
(Iii) Preparation of composition After mixing the cherry extract and sake lees extract at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0.34). %).
製造例3.組成物(3)の調製
(i)サクラの抽出物の調製
サトザクラの花を乾燥、粉砕し、粉砕物18gに精製水270gと1,3−ブチレングリコール630gを加え、80℃にて3時間抽出後、濾過し、褐色透明のサクラ抽出物溶液1350gを得た(固形分濃度0.39%)。
(ii)微生物由来発酵代謝物の調製
乳酸菌をMRS培地で、37℃で72時間液体培養し、培養液を濾過して得られる培養液を乳酸菌抽出物溶液(固形分濃度0.72%)とした。
(iii)組成物の調製
サクラ抽出物溶液と乳酸菌抽出物溶液を、1:2の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.51%)
Production example 3. Preparation of composition (3) (i) Preparation of cherry extract The flowers of Satozakura are dried and crushed, 270 g of purified water and 630 g of 1,3-butylene glycol are added to 18 g of the crushed product, and the mixture is extracted at 80 ° C. for 3 hours. Then, it was filtered to obtain 1350 g of a brown transparent cherry extract solution (solid content concentration: 0.39%).
(Ii) Preparation of fermentation metabolites derived from microorganisms Lactobacillus was liquid-cultured in MRS medium at 37 ° C. for 72 hours, and the culture solution obtained by filtering the culture solution was mixed with a lactic acid bacteria extract solution (solid content concentration 0.72%). did.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 51%)
製造例4.組成物(4)の調製
(i)サクラの抽出物の調製
サトザクラに代えてヤマザクラを用いるほかは、製造例1の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1350gを得た(固形分濃度0.43%)。
(ii)微生物由来発酵代謝物の調製
製造例1の(ii)と同様の方法で、酵母抽出物溶液を得た。
(iii)組成物の調製
サクラ抽出物溶液と酵母抽出物溶液を、1:1の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.37%)
Production example 4. Preparation of Composition (4) (i) Preparation of Sakura Extract 1350 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 1 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.43%).
(Ii) Preparation of microbial-derived fermented metabolites A yeast extract solution was obtained in the same manner as in (ii) of Production Example 1.
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 37%)
製造例5.組成物(5)の調製
(i)サクラの抽出物の調製
サトザクラに代えてヤマザクラを用いるほかは、製造例2の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1355gを得た(固形分濃度0.41%)。
(ii)微生物由来発酵代謝物の調製
製造例2の(ii)と同様の方法により酒粕抽出物溶液を得た。
(iii)組成物の調製
サクラの抽出物溶液と酒粕抽出物溶液を、1:9の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.32%)。
Production example 5. Preparation of Composition (5) (i) Preparation of Sakura Extract 1355 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 2 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.41%).
(Ii) Preparation of microbial-derived fermented metabolite A sake lees extract solution was obtained by the same method as in (ii) of Production Example 2.
(Iii) Preparation of composition After mixing the cherry extract solution and the sake lees extract solution at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0). .32%).
製造例6.組成物(6)の調製
(i)サクラの抽出物の調製
サトザクラに代えてヤマザクラを用いるほかは、製造例3の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1345gを得た(固形分濃度0.35%)。
(ii)微生物由来発酵代謝物の調製
製造例3の(iii)と同様の方法にて、乳酸菌抽出物溶液を得た。
(iii)組成物の調製
サクラ抽出物溶液と乳酸菌抽出物溶液を、1:2の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.49%)
Production example 6. Preparation of Composition (6) (i) Preparation of Sakura Extract 1345 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 3 (i) except that wild cherry was used instead of Satozakura. (Solid content concentration 0.35%).
(Ii) Preparation of microbial-derived fermented metabolite A lactic acid bacterium extract solution was obtained by the same method as in (iii) of Production Example 3.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 49%)
製造例7.組成物(7)の調製
(i)サクラの抽出物の調製
サトザクラに代えてソメイヨシノ(Prunus×yedoensis)を用いるほかは、製造例1の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1360gを得た(固形分濃度0.42%)。
(ii)微生物由来発酵代謝物の調製
製造例1の(ii)と同様の方法で、酵母抽出物溶液を得た。
(iii)組成物の調製
サクラ抽出物溶液と酵母抽出物溶液を、1:1の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.38%)
Production example 7. Preparation of composition (7) (i) Preparation of sakura extract A brown transparent sakura extract was prepared by the same method as in Production Example 1 (i) except that Yoshino cherry tree (Prunus × yedoensis) was used instead of Satozakura. 1360 g of the solution was obtained (solid content concentration 0.42%).
(Ii) Preparation of microbial-derived fermented metabolites A yeast extract solution was obtained in the same manner as in (ii) of Production Example 1.
(Iii) Preparation of composition After mixing the cherry extract solution and the yeast extract solution at a mixing ratio of 1: 1, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 38%)
製造例8.組成物(8)の調製
(i)サクラの抽出物の調製
サトザクラに代えてソメイヨシノを用いるほかは、製造例2の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1355gを得た(固形分濃度0.40%)。
(ii)微生物由来発酵代謝物の調製
製造例2の(ii)と同様の方法により酒粕抽出物溶液を得た。
(iii)組成物の調製
サクラの抽出物溶液と酒粕抽出物溶液を、1:9の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.31%)。
Production example 8. Preparation of Composition (8) (i) Preparation of Sakura Extract 1355 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 2 (i) except that Yoshino cherry tree was used instead of Satozakura. (Solid content concentration 0.40%).
(Ii) Preparation of microbial-derived fermented metabolite A sake lees extract solution was obtained by the same method as in (ii) of Production Example 2.
(Iii) Preparation of composition After mixing the cherry extract solution and the sake lees extract solution at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0). .31%).
製造例9.組成物(9)の調製
(i)サクラの抽出物の調製
サトザクラに代えてソメイヨシノを用いるほかは、製造例3の(i)と同様の方法により、褐色透明のサクラ抽出物溶液1340gを得た(固形分濃度0.34%)。
(ii)微生物由来発酵代謝物の調製
製造例3の(iii)と同様の方法にて、乳酸菌抽出物溶液を得た。
(iii)組成物の調製
サクラ抽出物溶液と乳酸菌抽出物溶液を、1:2の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.48%)
Production example 9. Preparation of Composition (9) (i) Preparation of Sakura Extract 1340 g of a brown transparent Sakura extract solution was obtained by the same method as in Production Example 3 (i) except that Yoshino cherry tree was used instead of Satozakura. (Solid content concentration 0.34%).
(Ii) Preparation of microbial-derived fermented metabolite A lactic acid bacterium extract solution was obtained by the same method as in (iii) of Production Example 3.
(Iii) Preparation of composition After mixing the cherry extract solution and the lactic acid bacterium extract solution at a mixing ratio of 1: 2, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 48%)
製造例10.組成物(10)の調製
(i)サクラの抽出物の調製
製造例2の(i)と同様の方法により、サクラ抽出物溶液を得た。
(ii)微生物由来発酵代謝物の調製
精白米50gを蒸して冷却した後、アスペルギルス オリゼー(Aspergillus oryzae)を添加し、これを30℃の温度条件で米麹を作製した。これに、別に蒸した精白米50gと殺菌した精製水200gを加え、さらに予め培養したサッカロミセス セレビシエ(Saccharomyces cerevisiae)培養液を添加して、30℃で発酵を行った。発酵終了後、この米発酵物溶液を90℃で1時間、加熱殺菌処理した後、室温に戻し、濾過をして淡黄色透明の米発酵物溶液126gを得た(固形分濃度4.00%)。
(iii)組成物の調製
サクラの抽出物と米発酵物溶液を、1:9の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.30%)。
Production example 10. Preparation of Composition (10) (i) Preparation of Sakura Extract A cherry extract solution was obtained by the same method as in Production Example 2 (i).
(Ii) Preparation of fermented metabolites derived from microorganisms After steaming and cooling 50 g of polished rice, Aspergillus oryzae was added to prepare rice jiuqu under a temperature condition of 30 ° C. To this, 50 g of separately steamed polished rice and 200 g of sterilized purified water were added, and a pre-cultured Saccharomyces cerevisiae culture solution was added, and fermentation was carried out at 30 ° C. After the fermentation was completed, this rice fermented product solution was heat sterilized at 90 ° C. for 1 hour, then returned to room temperature and filtered to obtain 126 g of a pale yellow transparent rice fermented product solution (solid content concentration: 4.00%). ).
(Iii) Preparation of composition After mixing the cherry extract and the fermented rice solution at a mixing ratio of 1: 9, the insoluble material was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0. 30%).
製造例11.組成物(11)の調製
(i)サクラの抽出物の調製
製造例3(i)と同様の方法により、サクラ抽出物溶液を得た。
(ii)微生物由来発酵代謝物の調製
米麹100gに予め滅菌しておいた精製水を300g加え、35℃で静置した。その後、これに別に蒸した精白米100gと予め培養したサッカロミセス セレビシエ(Saccharomyces cerevisiae)培養液を添加して30℃で発酵を行った。発酵終了後、この米発酵物溶液を90℃で1時間、加熱殺菌処理した後、室温に戻し、濾過をして淡黄色透明の米発酵物溶液183gを得た(固形分濃度3.93%)。
(iii)組成物の調製
サクラの抽出物と酒粕抽出物を、1:9の混合比で混合後、不溶物を濾過し、褐色透明の抽出物混合溶液を得た(固形分濃度0.31%)。
Production example 11. Preparation of Composition (11) (i) Preparation of Sakura Extract A cherry extract solution was obtained by the same method as in Production Example 3 (i).
(Ii) Preparation of fermented metabolites derived from microorganisms 300 g of purified water sterilized in advance was added to 100 g of rice jiuqu, and the mixture was allowed to stand at 35 ° C. Then, 100 g of separately steamed polished rice and a pre-cultured Saccharomyces cerevisiae culture solution were added thereto, and fermentation was carried out at 30 ° C. After the fermentation was completed, this rice fermented product solution was heat sterilized at 90 ° C. for 1 hour, then returned to room temperature and filtered to obtain 183 g of a pale yellow transparent rice fermented product solution (solid content concentration 3.93%). ).
(Iii) Preparation of composition After mixing the cherry extract and sake lees extract at a mixing ratio of 1: 9, the insoluble matter was filtered to obtain a brown transparent extract mixed solution (solid content concentration 0.31). %).
比較試料(1)の調製
製造例1の(i)と同様の方法によりサクラ抽出物溶液を調製し、これを比較試料(1)とした。
Preparation of Comparative Sample (1) A cherry extract solution was prepared by the same method as in Production Example 1 (i), and this was used as the comparative sample (1).
比較試料(2)の調製
製造例2の(ii)と同様の方法により酒粕抽出物溶液を調製し、これを比較試料(2)とした。
Preparation of Comparative Sample (2) A sake lees extract solution was prepared by the same method as in Production Example 2 (ii), and this was used as the comparative sample (2).
処方例1.化粧水
[A成分] 部
オリーブ油 1.0
ポリオキシエチレン(5.5)セチルアルコール 5.0
ブチルパラベン 0.1
[B成分]
製造例1の組成物(1) 5.0
エタノール 5.0
グリセリン 5.0
1,3−ブチレングリコール 5.0
水酸化カリウム 適量
精製水 全量が100部となる量
[C成分]
香料 適量
Prescription example 1. Toner [A component] Part Olive oil 1.0
Polyoxyethylene (5.5) cetyl alcohol 5.0
Butylparaben 0.1
[B component]
Composition of Production Example 1 (1) 5.0
Ethanol 5.0
Glycerin 5.0
1,3-butylene glycol 5.0
Potassium hydroxide Appropriate amount Purified water 100 parts in total [C component]
Appropriate amount of fragrance
処方例2.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例2の組成物(2)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 2. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (2) of Production Example 2 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例3.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例3の組成物(3)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 3. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (3) of Production Example 3 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例4.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例4の組成物(4)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 4. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (4) of Production Example 4 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例5.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例5の組成物(5)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 5. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (5) of Production Example 5 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例6.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例6の組成物(6)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 6. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (6) of Production Example 6 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例7.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例7の組成物(7)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 7. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (7) of Production Example 7 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例8.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例8の組成物(8)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 8. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (8) of Production Example 8 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例9.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例9の組成物(9)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 9. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (9) of Production Example 9 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例10.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例10の組成物(10)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 10. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (10) of Production Example 10 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例11.化粧水
処方例1のB成分中、製造例1の組成物(1)に代えて、製造例11の組成物(10)5.0部を用いるほかは処方例1と同様にして化粧水を得た。
Prescription example 11. Toner In the B component of Formulation Example 1, 5.0 parts of the composition (10) of Production Example 11 is used instead of the composition (1) of Production Example 1, and the lotion is prepared in the same manner as in Formulation Example 1. Obtained.
処方例12.乳液
[A成分] 部
流動パラフィン 6.0
ヘキサラン 4.0
ホホバ油 1.0
ポリオキシエチレン(20)ソルビタンモノステアレート 2.0
大豆レシチン 1.5
[B成分]
製造例2の組成物(2) 3.0
L−アスコルビン酸−2−グルコシド 2.0
水酸化カリウム 0.5
グリセリン 3.0
1,3−ブチレングリコール 2.0
カルボキシメチルセルロース 0.3
ヒアルロン酸ナトリウム 0.01
精製水 全量が100部となる量
[C成分]
香料 適量
Prescription example 12. Emulsion [Component A] Part Liquid paraffin 6.0
Hexalan 4.0
Jojoba oil 1.0
Polyoxyethylene (20) sorbitan monostearate 2.0
Soy lecithin 1.5
[B component]
Composition of Production Example 2 (2) 3.0
L-ascorbic acid-2-glucoside 2.0
Potassium hydroxide 0.5
Glycerin 3.0
1,3-butylene glycol 2.0
Carboxymethyl cellulose 0.3
Sodium hyaluronate 0.01
Amount of purified water totaling 100 parts [C component]
Appropriate amount of fragrance
処方例13.乳液
処方例12のB成分中、L−アスコルビン酸−2−グルコシド2.0部及び水酸化カリウム0.5部に代えてトラネキサム酸2.0部を用いるほかは処方例12と同様にして乳液を得た。
Prescription example 13. Emulsion Emulsion in the same manner as in Formulation 12, except that 2.0 parts of tranexamic acid is used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 part of potassium hydroxide in the B component of Formulation Example 12. Got
処方例14.乳液
処方例12のB成分中、L−アスコルビン酸−2−グルコシド2.0部及び水酸化カリウム0.5部に代えてアルブチン3.0部を用いるほかは処方例12と同様にして乳液を得た。
Prescription example 14. Emulsion Emulsion was prepared in the same manner as in Formulation 12 except that 3.0 parts of arbutin was used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 part of potassium hydroxide in the B component of Formulation Example 12. Obtained.
処方例15.乳液
処方例12のB成分中、L−アスコルビン酸−2−グルコシド2.0部及び水酸化カリウム0.5部に代えてニコチン酸アミド5.0部を用いるほかは処方例12と同様にして乳液を得た。
Prescription example 15. Emulsion Same as in Formulation 12 except that 5.0 parts of nicotinamide is used instead of 2.0 parts of L-ascorbic acid-2-glucoside and 0.5 parts of potassium hydroxide in the B component of Formulation Example 12. Emulsion was obtained.
処方例16.エッセンス
[成分] 部
エタノール 2.0
グリセリン 5.0
1,3−ブチレングリコール 5.0
メチルパラベン 0.1
ヒアルロン酸 0.1
製造例5の組成物(5) 5.0
クエン酸 0.3
クエン酸ナトリウム 0.6
精製水 全量が100部となる量
Prescription example 16. Essence [Ingredients] Part Ethanol 2.0
Glycerin 5.0
1,3-butylene glycol 5.0
Methylparaben 0.1
Hyaluronic acid 0.1
Composition of Production Example 5 (5) 5.0
Citric acid 0.3
Sodium citrate 0.6
Amount of purified water totaling 100 parts
処方例17.ローション
[成分] 部
製造例6の組成物(6) 10.0
エタノール 10.0
グリセリン 3.0
1、3−ブチレングリコール 2.0
メチルパラベン 0.2
クエン酸 0.1
クエン酸ナトリウム 0.3
カルボキシビニルポリマー 0.1
キサンタンガム 0.1
香料 適量
水酸化カリウム 適量
精製水 全量が100部となる量
上記の成分を混合してローションを得た。
Prescription example 17. Lotion [Ingredients] Part Composition of Production Example 6 (6) 10.0
Ethanol 10.0
Glycerin 3.0
1,3-butylene glycol 2.0
Methylparaben 0.2
Citric acid 0.1
Sodium citrate 0.3
Carboxyvinyl polymer 0.1
Xanthan gum 0.1
Perfume Appropriate amount Potassium hydroxide Appropriate amount Purified water 100 parts in total The above components were mixed to obtain a lotion.
実施例18.リキッドファンデーション
[A成分] 部
ステアリン酸 2.4
モノステアリン酸プロピレングリコール 2.0
セトステアリルアルコール 0.2
液状ラノリン 2.0
流動パラフィン 3.0
ミリスチン酸イソプロピル 8.5
プロピルパラベン 0.05
[B成分]
製造例7の組成物(7) 5.0
カルボキシメチルセルロースナトリウム 0.2
ベントナイト 0.5
プロピレングリコール 4.0
トリエタノールアミン 1.1
メチルパラベン 0.1
精製水 全量が100部となる量
[C成分]
酸化チタン 8.0
タルク 4.0
着色顔料 適量
Example 18. Liquid foundation [A component] Stearic acid 2.4
Propylene glycol monostearate 2.0
Setostearyl alcohol 0.2
Liquid lanolin 2.0
Liquid paraffin 3.0
Isopropyl myristate 8.5
Propylparaben 0.05
[B component]
Composition of Production Example 7 (7) 5.0
Sodium Carboxymethyl Cellulose 0.2
Bentonite 0.5
Propylene glycol 4.0
Triethanolamine 1.1
Methylparaben 0.1
Amount of purified water totaling 100 parts [C component]
Titanium oxide 8.0
Talc 4.0
Appropriate amount of coloring pigment
処方例19.ヘアシャンプー
[A成分] 部
N−ヤシ油脂肪酸メチルタウリンナトリウム 10.0
ポリオキシエチレン(3)アルキルエーテル硫酸ナトリウム 20.0
ラウリルジメチルアミノ酢酸ベタイン 10.0
ヤシ油脂肪酸ジエタノールアミド 4.0
メチルパラベン 0.1
[B成分]
クエン酸 0.1
製造例10の組成物(10) 2.0
1,3−ブチレングリコー ル 2.0
精製水 全量が100部となる量
Prescription example 19. Hair shampoo [Ingredient A] Part N-coconut oil fatty acid methyl taurine sodium 10.0
Polyoxyethylene (3) Alkyl Ether Sodium Sulfate 20.0
Betaine Lauryl Dimethylamino Acetate 10.0
Coconut oil fatty acid diethanolamide 4.0
Methylparaben 0.1
[B component]
Citric acid 0.1
Composition of Production Example 10 (10) 2.0
1,3-butylene glycol 2.0
Amount of purified water totaling 100 parts
実施例20.ヘアコンディショナー
[A成分] 部
ポリオキシエチレン(10)硬化ヒマシ油 1.0
塩化ジステアリルジメチルアンモニウム 1.5
塩化ステアリルトリメチルアンモニウム 2.0
2−エチルヘキサン酸グリセリル 1.0
セタノール 3.2
ステアリルアルコール 1.0
メチルパラベン 0.1
[B成分]
製造例1の組成物(1) 2.0
1,3−ブチレングリコール 5.0
精製水 全量が100部となる量
Example 20. Hair conditioner [Component A] Part Polyoxyethylene (10) Hardened castor oil 1.0
Distearyl dimethylammonium chloride 1.5
Stearyltrimethylammonium chloride 2.0
Glyceryl 2-ethylhexanoate 1.0
Cetanol 3.2
Stearyl alcohol 1.0
Methylparaben 0.1
[B component]
Composition of Production Example 1 (1) 2.0
1,3-butylene glycol 5.0
Amount of purified water totaling 100 parts
処方例21.育毛用ヘアトニック
[成分] 部
グリチルリチン酸ジカリウム 0.1
モノニトログアヤコールナトリウム 0.02
塩酸ピリドキシン 0.03
アデノシン 1.0
製造例2の組成物(2) 1.0
トリメチルグリシン 0.5
乳酸 0.2
1,3−ブチレングリコール 10.0
フェノキシエタノール 0.2
ポリオキシエチレン硬化ヒマシ油 0.4
タマサキツヅラフジ根エキス 0.3
オタネニンジンエキス 0.3
L−アルギニン 適量
精製水 全量が100部となる量
Prescription example 21. Hair tonic for hair growth [Ingredients] Part Dipotassium glycyrrhizinate 0.1
Mononitroguaiacol Sodium 0.02
Pyridoxine hydrochloride 0.03
Adenosine 1.0
Composition of Production Example 2 (2) 1.0
Trimethylglycine 0.5
Lactic acid 0.2
1,3-butylene glycol 10.0
Phenoxyethanol 0.2
Polyoxyethylene hydrogenated castor oil 0.4
Menispermaceae root extract 0.3
Panax ginseng extract 0.3
Appropriate amount of L-arginine Amount of purified water totaling 100 parts
本発明の組成物の有効性について以下の試験例1〜7により評価を行ったが、本発明はこれに限るものではない。 The effectiveness of the composition of the present invention was evaluated by the following Test Examples 1 to 7, but the present invention is not limited to this.
試験例1.表皮細胞賦活作用の評価
ヒト表皮細胞NHEKを、HuMedia KG2培地(クラボウ社製)を入れた96穴マイクロプレートに5×103個/穴播種し、37℃、5.0%CO2の条件下に1日間プレ培養した後、製造例1〜10の組成物(1)〜(10)のそれぞれを試料溶液として含む培地(Humedia KG2)をプレ培養液に添加し、同条件でさらに2日間培養した。ここで、試料溶液の濃度は、追添加する培地全量に対する溶液としての終濃度が1.0%、2.0%となるように調製した。プレ培養後、培地を除去し、0.03%のMTTを添加して37℃に1時間保持した後、生成したホルマザンをイソプロパノールで抽出し、マイクロプレートリーダー(Model 680、バイオラッド社製)を用いて波長570−630nmでMTT値を測定した。また、試料無添加の場合(Control)についても上記と同様の操作を行った。そして、試料無添加時のMTT値を100としたときの各試料添加時のMTT値の相対値を求め、表皮細胞MTT活性率(%)とした。また、試験系が正常に機能しているかを確認するために、試料溶液の代わりに陽性対照として100mMのグルコースを添加した場合についても、同様の試験を行った。
Test example 1. Evaluation of epidermal cell activating effect Human epidermal cell NHEK was seeded in 5 × 10 3 cells / hole in a 96-well microplate containing HuMedia KG2 medium (manufactured by Kurabo), and 1 under the conditions of 37 ° C and 5.0% CO 2. After pre-culturing for 1 day, a medium (Humedia KG2) containing each of the compositions (1) to (10) of Production Examples 1 to 10 as a sample solution was added to the pre-culture solution, and the cells were cultured under the same conditions for another 2 days. Here, the concentration of the sample solution was adjusted so that the final concentration as a solution with respect to the total amount of the medium to be added was 1.0% and 2.0%. After pre-culture, the medium is removed, 0.03% MTT is added and the temperature is kept at 37 ° C. for 1 hour, and then the produced formazan is extracted with isopropanol and used with a microplate reader (Model 680, manufactured by Biorad). The MTT value was measured at a wavelength of 570-630 nm. In addition, the same operation as above was performed in the case of no sample addition (Control). Then, the relative value of the MTT value at the time of adding each sample was obtained when the MTT value at the time of no sample addition was 100, and used as the epidermal cell MTT activity rate (%). A similar test was also performed when 100 mM glucose was added as a positive control instead of the sample solution to confirm that the test system was functioning normally.
試験例1の結果を表1に示す。
[表1]
The results of Test Example 1 are shown in Table 1.
[Table 1]
表1に示すように、本発明に係る組成物(1)〜(10)は、濃度依存的に格段にすぐれた表皮細胞賦活効果を有することが確認された。また、グルコースにおいても、同様の効果が得られたことから、本試験系が正常に機能していることも確認された。 As shown in Table 1, it was confirmed that the compositions (1) to (10) according to the present invention have a remarkably excellent epidermal cell activating effect in a concentration-dependent manner. In addition, the same effect was obtained with glucose, confirming that this test system is functioning normally.
試験例2.DPPHラジカル消去作用の評価
DPPH(1,1−ジフェニル−2−ピクリルヒドラジル)2.4部をエタノール20部に溶解し、これに精製水20部を加えてDPPH溶液を調製した。このDPPH溶液24部に対して、18v/v%エタノール溶液を19.2部、2M酢酸−酢酸ナトリウム緩衝液(pH5.5)を4.8部加えて、DPPH添加溶液として調製した。また、抽出液そのものの色調が試験に及ぼす影響を差し引くため、DPPH溶液の代わりに50v/v%エタノール溶液を用いて、18v/v%エタノール溶液と2M酢酸−酢酸ナトリウム緩衝液を混合した液を対照液とした。次に、製造例1〜10の組成物(1)〜(10)及び比較試料(1)〜(2)を精製水でそれぞれ希釈して12種の試料溶液を調製した。なお、各組成物は、試料溶液全量に対する溶液としての終濃度がそれぞれ1.0%、2.0%となるように精製水で希釈した。この試料溶液とDPPH添加溶液又は対照液とを1:3の割合で混合し、室温で10分静置後、各試料溶液をDPPH添加溶液と混合した場合の550nmにおける吸光度と、各試料溶液を対照液と混合した場合の550nmにおける吸光度との差を測定し、DPPHラジカルの残存量を確認した。また、試料無添加の場合(Control)についても上記と同様の操作を行い、試料無添加時のDPPHラジカル残存率を100としたときの各試料添加時のDPPHラジカル残存率の相対値を求めた。また、試験系が正常に機能しているかを確認するために、試料溶液の代わりに陽性対照として水溶性ビタミンE[Trolox](終濃度25μM)を添加した場合についても、同様の試験を行った。
Test example 2. Evaluation of DPPH radical scavenging effect
2.4 parts of DPPH (1,1-diphenyl-2-picrylhydrazil) was dissolved in 20 parts of ethanol, and 20 parts of purified water was added thereto to prepare a DPPH solution. To 24 parts of this DPPH solution, 19.2 parts of 18v / v% ethanol solution and 4.8 parts of 2M sodium acetate-sodium acetate buffer (pH 5.5) were added to prepare a DPPH-added solution. In addition, in order to subtract the effect of the color tone of the extract itself on the test, a 50v / v% ethanol solution was used instead of the DPPH solution, and a mixture of 18v / v% ethanol solution and 2M sodium acetate-sodium acetate buffer was used. It was used as a control solution. Next, the compositions (1) to (10) of Production Examples 1 to 10 and the comparative samples (1) to (2) were diluted with purified water to prepare 12 kinds of sample solutions. Each composition was diluted with purified water so that the final concentration as a solution with respect to the total amount of the sample solution was 1.0% and 2.0%, respectively. This sample solution is mixed with the DPPH-added solution or the control solution at a ratio of 1: 3, and after allowing to stand at room temperature for 10 minutes, the absorbance at 550 nm when each sample solution is mixed with the DPPH-added solution and each sample solution are measured. The difference from the absorbance at 550 nm when mixed with the control solution was measured to confirm the residual amount of DPPH radicals. In addition, the same operation as above was performed in the case of no sample addition (Control), and the relative value of the DPPH radical residual rate at the time of adding each sample was obtained when the DPPH radical residual rate at the time of no sample addition was 100. .. A similar test was also performed when water-soluble vitamin E [Trolox] (final concentration 25 μM) was added as a positive control instead of the sample solution to confirm that the test system was functioning normally. ..
試験例2の結果を表2に示す。
[表2]
The results of Test Example 2 are shown in Table 2.
[Table 2]
表2に示すように、本発明に係る組成物(1)〜(10)は、比較試料(1)及び(2)と比較して、濃度依存的に格段にすぐれたDPPHラジカル消去作用を有することが示された。また、水溶性ビタミンE[Trolox]においても、同様の効果が得られたことから、本試験系が正常に機能していることも確認された。 As shown in Table 2, the compositions (1) to (10) according to the present invention have a concentration-dependently excellent DPPH radical scavenging action as compared with the comparative samples (1) and (2). Was shown. In addition, the same effect was obtained with water-soluble vitamin E [Trolox], confirming that this test system is functioning normally.
試験例3.SOD様作用の評価
1Mトリス−塩酸緩衝液0.15mL、1mMエチレンジアミン四酢酸・二ナトリウム塩溶液0.30mL、1mMキサンチン溶液0.30mL、0.75mMニトロブル-テトラゾリウム溶液0.20mL、製造例1〜10の組成物(1)〜(10)及び比較試料(1)〜(2)の各0.10mLと精製水1.90mLとを混合して12種の試験溶液を調製した。また、試験溶液に代えて精製水2.00mLを用いる他は上記試験溶液と同様の組成からなる混合液(コントロール[Control])を調製した。さらに、試料溶液(0.10mL)に代えて、0.875Unit/mLのスーパーオキシドジスムターゼ(SOD)溶液0.10mLを用いる他は上記試験溶液と同様の組成からなる混合液(陽性対照液)を調製した。上記試験溶液をそれぞれ37℃でインキュベートした後、これに1Unit/mLキサンチンオキシダーゼ溶液0.05mLを添加し、一定時間経過後(5分)、各試験溶液の570nmでの吸光度(被験液中のスーパーオキシドアニオン量の指標)を測定した。測定結果は、試料無添加(Control)の混合液の吸光度を100とした時の各試験溶液及び陽性対照液の吸光度を相対値で示した。
Test example 3. Evaluation of SOD-like action
1M Tris-hydrochloric acid buffer 0.15 mL, 1 mM ethylenediamine tetraacetic acid / disodium salt solution 0.30 mL, 1 mM xanthin solution 0.30 mL, 0.75 mM nitrobul-tetrazolium solution 0.20 mL, compositions of Production Examples 1 to 10 (1) to (10) ) And 0.10 mL of each of the comparative samples (1) and (2) and 1.90 mL of purified water were mixed to prepare 12 kinds of test solutions. In addition, a mixture (Control] having the same composition as the above test solution was prepared except that 2.00 mL of purified water was used instead of the test solution. Further, a mixed solution (positive control solution) having the same composition as the above test solution was prepared except that 0.10 mL of 0.875 Unit / mL superoxide dismutase (SOD) solution was used instead of the sample solution (0.10 mL). After incubating each of the above test solutions at 37 ° C, 0.05 mL of 1 Unit / mL xanthine oxidase solution was added thereto, and after a certain period of time (5 minutes), the absorbance of each test solution at 570 nm (superoxide in the test solution). An index of the amount of anion) was measured. The measurement results showed the absorbances of each test solution and positive control solution as relative values when the absorbance of the sample-free (Control) mixture was 100.
試験例3の結果を表3に示す。
[表3]
The results of Test Example 3 are shown in Table 3.
[Table 3]
表3に示すように、本発明に係る組成物(1)〜(10)は、比較試料(1)及び(2)と比較して、濃度依存的に格段にすぐれたSOD様活性を有することが示された。また、陽性対照であるSODにおいても、同様の効果が得られたことから、本試験系が正常に機能していることも確認された。 As shown in Table 3, the compositions (1) to (10) according to the present invention have significantly superior SOD-like activity in a concentration-dependent manner as compared with the comparative samples (1) and (2). It has been shown. In addition, the same effect was obtained in SOD, which is a positive control, confirming that this test system is functioning normally.
試験例4.XVII型コラーゲン遺伝子発現の評価
正常ヒト表皮細胞を、増殖添加剤含有HuMedia KG2培地(クラボウ社製)にて6×105個/mLに調製し、φ6cmシャーレに1mLを播種して、5%CO2、飽和水蒸気下、37℃で培養した。24時間培養後、さらに、製造例1〜3及び10の組成物(1)〜(3)及び(10)を試料溶液として含んだ同培地(培地全量に対して溶液としての終濃度が1.0%となるように試料溶液が含まれるもの)を追添加して培養した。また、試料無添加(Control)の場合についても上記と同様の操作を行った。24時間培養後、それぞれの試験区の細胞をTrizol試薬(Invitrogen社製)1mLにより回収した。回収した細胞に対してクロロホルム(和光純薬工業社製)200μLを添加して撹拌混合し、遠心分離機(TOMY社製/MX-160)で15,000rpm、4℃の条件下で15分間遠心分離した後、水層のみを400μL分取した。回収した水層にイソプロパノール(和光純薬工業社製)500μLを添加して撹拌混合し、15,000rpm、4℃の条件下で15分間遠心分離してtotal RNAの沈殿物を得た。total RNAに75%エタノールを1mL添加して撹拌して洗浄し、15,000rpm、4℃条件下で15分間遠心分離して沈殿を回収した。回収したtotal RNAを所定のキット(PrimeScript RT reagent Kit with gDNA Eraser (Perfect Real Time)[タカラバイオ社製])を用いて逆転写反応し、cDNAを合成した。合成したcDNAをサンプルとして、Thermal Cycler Dice(登録商標)Real Time System Single(タカラバイオ社製)、及びSYBR(登録商標)Premix Ex TaqTM II(Perfect Real Time)[タカラバイオ社製]を用いて、XVII型コラーゲン遺伝子の発現と、内部標準物質βアクチン遺伝子の発現の検出を行った。ここで、βアクチンは、ハウスキーピング遺伝子(多くの組織や細胞中に共通して一定量発現する遺伝子であって、常に発現され,細胞の維持,増殖に不可欠な遺伝子である)の一つであり、発現量が常に一定とされていることから、PCRの実験では内部標準として用いられるものである。試験結果は、βアクチン遺伝子の発現量を一定とした場合の、それぞれの試験区でのXVII型コラーゲン遺伝子の発現量を比較した。本試験系においては、コントロール区のそれぞれの遺伝子の発現量を100としたときの試験区での当該遺伝子の発現量の相対値を求めた。
Test example 4. Evaluation of XVII type collagen gene expression Normal human epidermal cells were prepared in 6 × 10 5 cells / mL in HuMedia KG2 medium (manufactured by Kurabo) containing a growth additive, and 1 mL was seeded in a φ6 cm petri dish to generate 5% CO. 2. Incubated at 37 ° C under saturated steam. After culturing for 24 hours, the same medium containing the compositions (1) to (3) and (10) of Production Examples 1 to 3 and 10 as a sample solution (the final concentration as a solution is 1.0% with respect to the total amount of the medium). (The one containing the sample solution) was added and cultured. In addition, the same operation as above was performed in the case of no sample addition (Control). After culturing for 24 hours, cells in each test group were collected with 1 mL of Trizol reagent (manufactured by Invitrogen). Add 200 μL of chloroform (manufactured by Wako Pure Chemical Industries, Ltd.) to the collected cells, stir and mix, and centrifuge with a centrifuge (manufactured by TOMY / MX-160) at 15,000 rpm for 15 minutes at 4 ° C. After that, only the aqueous layer was separated by 400 μL. 500 μL of isopropanol (manufactured by Wako Pure Chemical Industries, Ltd.) was added to the recovered aqueous layer, stirred and mixed, and centrifuged at 15,000 rpm and 4 ° C. for 15 minutes to obtain a total RNA precipitate. 1 mL of 75% ethanol was added to total RNA, stirred and washed, and centrifuged at 15,000 rpm for 15 minutes at 4 ° C. to recover the precipitate. The collected total RNA was reverse-transcribed using a predetermined kit (PrimeScript RT reagent Kit with gDNA Eraser (Perfect Real Time) [manufactured by Takara Bio Inc.]) to synthesize cDNA. Using the synthesized cDNA as a sample, using Thermal Cycler Dice (registered trademark) Real Time System Single (manufactured by Takara Bio) and SYBR (registered trademark) Premix Ex TaqTM II (Perfect Real Time) [manufactured by Takara Bio]. The expression of the XVII type collagen gene and the expression of the internal standard β-actin gene were detected. Here, β-actin is one of the housekeeping genes (genes that are commonly expressed in many tissues and cells in a certain amount, are always expressed, and are indispensable for cell maintenance and proliferation). Since the expression level is always constant, it is used as an internal standard in PCR experiments. The test results compared the expression levels of the XVII collagen gene in each test group when the expression level of the β-actin gene was constant. In this test system, the relative value of the expression level of the gene in the test group was determined when the expression level of each gene in the control group was 100.
試験例4の結果を表4に示す。
[表4]
The results of Test Example 4 are shown in Table 4.
[Table 4]
表4に示すように、本発明に係る組成物は、XVII型コラーゲンの遺伝子発現を顕著に誘導することが示された。 As shown in Table 4, the composition according to the present invention was shown to significantly induce gene expression of type XVII collagen.
試験例5.XVII型コラーゲン合成促進効果
正常ヒト皮膚由来表皮細胞(NHEK)をHuMedia KG2培地(クラボウ社製)を入れた96穴マイクロプレートに8×103個/穴播種し、37℃,5.0%CO2の条件下に1日間プレ培養した後、試料溶液を含む同培地(培地全量に対して溶液としての終濃度が1.0%,2.0%となるように試料溶液が含まれるもの)を追添加し、同条件でさらに2日間培養した。その後XVII型コラーゲン抗体を用いた免疫的検出を行った。すなわち、PBS(-)洗浄後、15%中性緩衝ホルマリン液を用いて細胞を30分処理して固定、0.5%Triton X-100溶液で1時間浸透処理、5倍希釈ブロッキングワンP(ナカライテスク社)溶液で2時間処理によるブロッキングを行った後、XVII型コラーゲン抗体を添加し、4℃で一昼夜静置した。その後PBS(-)洗浄し、蛍光ラベルした二次抗体を添加してさらに暗所で一定時間静置した。そのPBS(-)後洗浄し、蛍光強度の測定を行った。まず、二次抗体の蛍光ラベル(Alexa Fluor488)をEx=485nm、Em=520nmで測定し[蛍光マイクロプレートリーダー(フルオロスキャンアセント、Thermo Fisher Scientific社製)]、その後、Hoechst33342によるDNA染色を行い、Ex=355nm、Em=460nmの測定を行った。それぞれの試験区のAlexa Fluor488の蛍光強度をHoechst33342の蛍光強度で割ることで、XVII型コラーゲンの生成度合いを求めた。また、試料無添加の場合(Control)についても上記と同様の操作を行い、ここに得られたXVII型コラーゲン生成度合いに対する各試料添加時のXVII型コラーゲン生成度合いの相対値を求め、XVII型コラーゲン生成量(%)とした。
Test example 5. XVII type collagen synthesis promoting effect Normal human skin-derived epidermal cells (NHEK) were seeded in 96-well microplates containing HuMedia KG2 medium (manufactured by Kurabo) at 8 × 10 3 cells / hole at 37 ° C, 5.0% CO 2 . After pre-culturing under the conditions for 1 day, the same medium containing the sample solution (the one containing the sample solution so that the final concentration as a solution is 1.0% or 2.0% with respect to the total amount of the medium) is added and added. The cells were cultured for another 2 days under the conditions. After that, immunological detection was performed using a type XVII collagen antibody. That is, after washing with PBS (-), cells were treated with 15% neutral buffered formalin solution for 30 minutes to fix them, permeated with 0.5% Triton X-100 solution for 1 hour, and 5-fold diluted blocking one P (Nacalai Tesque). After blocking by treatment with a solution for 2 hours, type XVII collagen antibody was added, and the mixture was allowed to stand at 4 ° C. for 24 hours. Then, it was washed with PBS (-), a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand in a dark place for a certain period of time. After the PBS (-), it was washed and the fluorescence intensity was measured. First, the fluorescent label (Alexa Fluor488) of the secondary antibody was measured at Ex = 485 nm and Em = 520 nm [fluorescent microplate reader (Fluoroscan Ascent, manufactured by Thermo Fisher Scientific)], and then DNA staining with Hoechst33342 was performed. Measurements were performed at Ex = 355 nm and Em = 460 nm. The degree of XVII collagen production was determined by dividing the fluorescence intensity of Alexa Fluor 488 in each test group by the fluorescence intensity of Hoechst 33342. In addition, in the case of no sample addition (Control), the same operation as above is performed, and the relative value of the XVII type collagen production degree at the time of adding each sample to the XVII type collagen production degree obtained here is obtained, and the XVII type collagen is obtained. The amount produced (%) was used.
試験例5の結果を表5に示す。
[表5]
The results of Test Example 5 are shown in Table 5.
[Table 5]
表5に示すように、本発明に係る組成物は、濃度依存的にすぐれたXVII型コラーゲン合成促進作用を有することが確認された。XVII型コラーゲンは表皮と真皮の接合部に存在する基底膜と表皮とを結合させる役割を果たし、基底膜は基底細胞の分化形質発現を維持する役割を果たすことから、本発明に係る組成物は、XVII型コラーゲンの発現を誘導することで、基底膜を正常な状態に維持し、又皮膚の新陳代謝(ターンオーバー)を正常な状態に維持する効果を有することが示唆される。また、頭皮のXVII型コラーゲンの発現を誘導することで、脱毛、白髪予防効果を奏することも示唆される。 As shown in Table 5, it was confirmed that the composition according to the present invention has an excellent concentration-dependent type XVII collagen synthesis promoting action. Type XVII collagen plays a role of binding the basement membrane and the epidermis existing at the junction between the epidermis and the dermis, and the basement membrane plays a role of maintaining the expression of differentiation traits of the basal cells. , It is suggested that by inducing the expression of type XVII collagen, it has the effect of maintaining the basement membrane in a normal state and the skin metabolism (turnover) in a normal state. It is also suggested that by inducing the expression of type XVII collagen in the scalp, it has an effect of preventing hair loss and gray hair.
試験例6.メラニン合成抑制効果
培養B16マウスメラノーマ細胞B16−F10を、24穴マイクロプレートに2.4×104個/穴播種し、10%FBS含有RPMI1640培地中、37℃、5.0%CO2の条件下に1日間プレ培養した後、同培地で、試料溶液を溶液として終濃度が1.0%、2.0%となるように希釈した溶液と終濃度1mMになるように調整したテオフィリン含有培地を添加し、同条件で2日間培養した。次に培養液を除去し、1N NaOH/10%ジメチルスルフォキシド溶液を1穴あたり200μL添加し、シールして50℃、2時間インキュベートして細胞を溶解させた。この溶液100μLを別の96穴マイクロプレートに移し、マイクロプレートリーダー(Model 680、バイオラッド社製)を用い、波長490nmでメラニン値を測定した。一方同じ細胞を溶解させた溶液を5μL別の96穴マイクロプレートに移し、さらに精製水で5倍希釈したDye Reagent Concentrate(バイオラッド社)溶液を200μL添加し、 マイクロプレートリーダー(Model 680、バイオラッド社製)を用い、波長570nmの吸光度を測定した。別で既知の量の牛血清アルブミン(Sigma社製)を段階希釈し、同様に操作して得られた検量線から、アルブミン当量のタンパク質量を計測した。得られた吸光度をタンパク質量で除算して、タンパク質あたりのメラニン量を求めた。また、試料無添加の場合(Control)についても上記と同様の操作を行い、ここに得られたタンパク質あたりのメラニン量に対する各試料添加時のタンパク質あたりのメラニン量の相対値を求め、メラニン合成率(%)とした。なお、比較のため、試料溶液の代わりに、2mMのコウジ酸を添加した場合(陽性対照)についても同様の試験を行った。
Test example 6. Suppressive effect on melanin synthesis Cultured B16 mouse melanoma cells B16-F10 were seeded in a 24-well microplate at 2.4 × 10 4 cells / well, and were seeded in RPMI1640 medium containing 10% FBS for 1 day under the conditions of 37 ° C and 5.0% CO 2. After pre-culture, the sample solution was used as a solution, diluted to a final concentration of 1.0% and 2.0%, and a theophylline-containing medium adjusted to a final concentration of 1 mM was added, and under the same conditions, 2 Incubated for days. The culture was then removed, 200 μL of 1N NaOH / 10% dimethylsulfoxide solution was added per hole, sealed and incubated at 50 ° C. for 2 hours to lyse the cells. 100 μL of this solution was transferred to another 96-well microplate, and the melanin level was measured at a wavelength of 490 nm using a microplate reader (Model 680, manufactured by Bio-Rad). On the other hand, 5 μL of the solution in which the same cells were lysed was transferred to another 96-well microplate, and 200 μL of Dye Reagent Concentrate (Biorad) solution diluted 5-fold with purified water was added, and a microplate reader (Model 680, Biorad) was added. The absorbance at a wavelength of 570 nm was measured using (manufactured by the same company). Another known amount of bovine serum albumin (manufactured by Sigma) was serially diluted, and the amount of protein equivalent to albumin was measured from the calibration curve obtained by the same operation. The obtained absorbance was divided by the amount of protein to determine the amount of melanin per protein. Further, in the case of no sample addition (Control), the same operation as above is performed, and the relative value of the melanin amount per protein at the time of adding each sample to the melanin amount per protein obtained here is obtained, and the melanin synthesis rate is obtained. It was set to (%). For comparison, the same test was performed when 2 mM kojic acid was added instead of the sample solution (positive control).
試験例6の結果を表6に示す。
[表6]
The results of Test Example 6 are shown in Table 6.
[Table 6]
表6に示す通り、本発明に係る組成物は、格段にすぐれたメラニン合成抑制作用を有するものであり、シミ、ソバカスの予防、改善効果が示唆される。 As shown in Table 6, the composition according to the present invention has a remarkably excellent melanin synthesis inhibitory effect, suggesting a preventive and ameliorating effect on spots and freckles.
試験例7.ペリリピン合成抑制
以下、本発明に係る組成物の脂肪蓄積の抑制効果を、脂肪細胞内の脂肪滴周辺に存在するタンパク質[ペリリピン1(perilipin 1)]の合成抑制効果により評価する。このペリリピン1が減少すると、脂肪滴数の減少及び脂肪滴サイズの縮小が生じることが知られていることから、ペリリピン1の合成抑制効果を評価する。
マウス線維芽細胞(3T3-L1)を、10%FBS含有ダルベッコ変法イーグル最少必須培地(DMEM:日水製薬株式会社)を入れた96穴マイクロプレートに1.5×104個/穴播種し、37℃,5.0%CO2の条件下に3日間プレ培養した後、分化誘導培地(10%FBS、0.5mM3-イソブチル-1-メチルキサンチン(IBMX)、0.25μMデキサメタソン(DEX)及び1.1μg/mLインスリンを混合したDMEM)を添加後、さらに2日間培養した。その後、試料溶液を含む同培地(培地全量に対して溶液としての終濃度が1.0%,2.0%となるように試料溶液が含まれるもの)を追添加し、同条件でさらに5日間培養した。その後、ペリリピン1(perilipin1)抗体を用いた免疫的検出を行った。すなわち、PBS(-)洗浄後、細胞を10%中性緩衝ホルマリン液にて30分処理して固定、0.5%Triton X-100溶液で1時間浸透処理、5倍希釈ブロッキングワンP(ナカライテスク社)溶液で2時間処理によるブロッキングを行った後、perilipin1抗体を添加し、室温で1時間静置した。その後PBS(-)洗浄し、蛍光ラベルした二次抗体を添加してさらに暗所で一定時間静置した。その後PBS(-)洗浄し、蛍光強度の測定を行った。まず、二次抗体の蛍光ラベル(Alexa Fluor488)をEx=485nm、Em=520nmで測定し[蛍光マイクロプレートリーダー(フルオロスキャンアセント、Thermo Fisher Scientific社製)]、その後、Hoechst33342によるDNA染色を行い、Ex=355nm、Em=460nmの測定を行った。それぞれの試験区のAlexa Fluor488の蛍光強度をHoechst33342の蛍光強度で割ることで、perilipin1の生成度合いを求めた。また、試料無添加の場合(Control)についても上記と同様の操作を行い、ここに得られたperilipin1生成度合いに対する各試料添加時のperilipin1生成度合いの相対値を求め、perilipin1合成率(%)とした。
Test example 7. Suppression of Perilipin Synthesis Hereinafter, the effect of suppressing fat accumulation of the composition according to the present invention will be evaluated based on the effect of suppressing the synthesis of the protein [perilipin 1] present around the lipid droplets in adipocytes. Since it is known that when the amount of perilipin 1 decreases, the number of fat droplets decreases and the size of lipid droplets decreases, the effect of suppressing the synthesis of perilipin 1 is evaluated.
Mouse fibroblasts (3T3-L1) were seeded at 1.5 x 10 4 cells / hole in a 96-well microplate containing 10% FBS-containing Dalveco modified Eagle's minimum essential medium (DMEM: Nissui Pharmaceutical Co., Ltd.), 37 After preculture for 3 days under the conditions of ℃ and 5.0% CO 2 , differentiation-inducing medium (10% FBS, 0.5 mM 3-isobutyl-1-methylxanthin (IBMX), 0.25 μM dexamethasone (DEX) and 1.1 μg / mL insulin) After adding DMEM) mixed with, the cells were cultured for another 2 days. Then, the same medium containing the sample solution (the one containing the sample solution so that the final concentration as a solution was 1.0% and 2.0% with respect to the total amount of the medium) was additionally added, and the cells were cultured under the same conditions for another 5 days. Then, immunodetection was performed using a perilipin1 antibody. That is, after washing with PBS (-), cells were treated with 10% neutral buffered formalin solution for 30 minutes to fix, permeated with 0.5% Triton X-100 solution for 1 hour, and 5-fold diluted Blocking One P (Nacalai Tesque). ) After blocking by treatment with the solution for 2 hours, perilipin1 antibody was added, and the mixture was allowed to stand at room temperature for 1 hour. Then, it was washed with PBS (-), a fluorescently labeled secondary antibody was added, and the mixture was allowed to stand in a dark place for a certain period of time. After that, it was washed with PBS (-) and the fluorescence intensity was measured. First, the fluorescent label (Alexa Fluor488) of the secondary antibody was measured at Ex = 485 nm and Em = 520 nm [fluorescent microplate reader (Fluoroscan Ascent, manufactured by Thermo Fisher Scientific)], and then DNA staining with Hoechst33342 was performed. Measurements were performed at Ex = 355 nm and Em = 460 nm. The degree of perilipin1 formation was determined by dividing the fluorescence intensity of Alexa Fluor 488 in each test group by the fluorescence intensity of Hoechst 33342. In addition, in the case of no sample addition (Control), the same operation as above was performed, and the relative value of the perilipin1 formation degree at each sample addition was obtained with respect to the perilipin1 formation degree obtained here, and the perilipin1 synthesis rate (%) was obtained. did.
試験例7の結果を表7に示す。
[表7]
The results of Test Example 7 are shown in Table 7.
[Table 7]
表7に示すように、本発明に係る組成物は、濃度依存的に格段にすぐれたペリリピン合成抑制効果を有することが確認された。 As shown in Table 7, it was confirmed that the composition according to the present invention has a remarkably excellent effect of suppressing perilipin synthesis in a concentration-dependent manner.
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